Stability of environmental DNA methylation and its utility in tracing spawning in fish.

The use of environmental DNA (eDNA) is becoming prevalent as a novel method of ecological monitoring. Although eDNA can provide critical information on the distribution and biomass of particular taxa, the DNA sequences of an organism remain unaltered throughout its existence, which complicates the accurate identification of crucial events, including spawning. Therefore, we examined DNA methylation as a novel source of information from eDNA, considering that the methylation patterns in eggs and sperm released during spawning differ from those of somatic tissues. Despite its potential applications, little is known about eDNA methylation, including its stability and methods for detection and quantification. Therefore, we conducted tank experiments and performed methylation analysis targeting 18S rDNA through bisulphite amplicon sequencing. In the target region, eDNA methylation was not affected by degradation and was equivalent to the methylation rate of genomic DNA from somatic tissues. Unmethylated DNA, abundant in the ovaries, was detected in the eDNA released during fish spawning. These results indicate that eDNA methylation is a stable signal reflecting targeted gene methylation and further demonstrate that germ cell-specific methylation patterns can be used as markers for detecting fish spawning.

Unveiling the effects of environmental factors and Yellow River inputs on the ichthyoplankton community structure in typical bays.

To unravel the effects of environmental factors on fishery resources in the bay, we conducted six biological and environmental surveys in the Laizhou Bay between 2013 and 2020. The findings of our study illuminated several key aspects: (1) The annual discharge of water and sediment from the Yellow River to Laizhou Bay exhibited notable variations, while concurrently, environmental factors including temperature, salinity, and suspended particle matter underwent fluctuations, yet remained within a relatively stable range overall. (2) A total of 8318 eggs and larvae belonging to 10 orders, 16 families, and 19 genera were collected. Significant interannual fluctuations had been documented in the species composition, abundance, and biodiversity of ichthyoplankton. Notably, both Shannon-Wiener diversity index and Pielou evenness index were significantly negatively correlated with suspended particle matter concentration. (3) The water and sediment discharge significantly positively correlated with the number of cold-temperature species. However, the sediment input negatively correlated with the number of continental shelf benthopelagic fish. (4) Redundancy and correlation analyses confirmed the strong link between spatial and temporal distribution of fish communities and environmental factors, with salinity and dissolved oxygen key for ichthyoplankton abundance. Our research offers a scientific foundation for targeted fishery protection and management, which is crucial for preserving the ecological functions of spawning grounds in the bay.

Study on the key ecological factors for the reproduction of Gymnocypris eckloni in the upper reaches of the Yellow River.

The development of hydroelectric projects has adversely affected the reproductive activities of downstream fish species. To facilitate the natural reproduction of fish and restore spawning grounds post-dam construction, it is imperative to explore the ecological factors crucial for their reproduction. Currently, various research methods with different advantages and limitations are employed for this purpose. Using identified spawning locations and periods as clues, we quantitatively investigate the flow velocity, water depth, water temperature, and riverbed substrate required for spawning. The results are validated using habitat simulation methods, aiming to establish a more scientific approach to explore ecological factors affecting fish reproduction. This study provides a more scientific, systematic, and detailed report on the ecological factors required for the spawning of Gymnocypris eckloni: flow velocity ranging from 0.19 to 0.97 m/s, water depth from 0.28 to 1.12 m, water temperature between 11.4 and 15.2°C, and predominantly gravel substrate. The reliability of the results was verified in another spawning ground, with good verification results. This research provides crucial data for the bio-mimetic reproductive technology of Gymnocypris eckloni and the restoration of spawning grounds for natural fish reproduction post-dam construction. It addresses the lack of suitable ecological factor data for protective fish species in the upper reaches of the Yellow River. The method exhibits strong scientific, accurate, and implementable characteristics.

Ovarian histology of the freshwater catfish Silonia silondia (Hamilton, 1822).

Histological characterization of gonadal tissues and their development can divulge vital clues to interpret the onset and progression of the spawning period in fish species and help in wild stock augmentation to conserve the natural fish stocks. In the present study, ovarian histology of female Silonia silonda was investigated from the Kangsha River, Bangladesh from March to November 2018. During this investigation, 54 females of Silonia silondia was analyzed to study the ovarian development. The histological evaluation revealed five distinct stages of gonadal development (Immature, Developing, Spawning capable, Regression or spent and Regeneration or Re-developing). Six phases of oocyte development were further characterized during the gonadal development stages as under-developed, primary growth, cortical alveolar, vitellogenic, pre-mature, and mature stage. In conclusion, the gonado-somatic index and histological analysis of ovaries indicated that the peak spawning of S. silondia occurs in July, and suggesting that the studied fish spawns once per breeding season. The present study recommends year round investigation of gonadal cycle and gonad developmental study of male individuals of S. silonida.

Tracking movements of meagre (Argyrosomus regius) during the spawning season: Preliminary indications of off estuary spawning.

Understanding the spawning behavior of meagre (Argyrosomus regius) is crucial for fisheries management and conservation. Meagre forms large spawning aggregations in estuaries, yet details of its spawning grounds remain elusive. We tagged 41 individuals and monitored their movements throughout several spawning seasons. Detections inside estuaries were limited. Fish detected inside the Tejo estuary during the spawning season performed regular movements toward coastal areas, suggesting potential spawning outside estuaries. Our findings underscore the significance of understanding meagre's migratory patterns for effective fisheries management and conservation strategies, supporting the establishment of spatial and seasonal closures during the spawning season.

Errors in estimating reproductive parameters with macroscopic methods: a case study on the protogynous blacktip grouper Epinephelus fasciatus (Forsskål 1775).

A size-based, histological analysis of the reproductive life history of the blacktip grouper, Epinephelus fasciatus (Forsskål 1775), was conducted in Indonesia to evaluate the error rate associated with macroscopic reproductive analysis. Histological results indicated that E. fasciatus was protogynous with female L50 at 13.4 cm total length (LT) and a size at sexual transition of 22.0 cm LT. The weight-length relationship for the species was W = 0.011 L3.13. Overall sex ratios were significantly female biased, operational sex ratios were significantly male biased, and sex ratios of mature individuals varied predictably with length from female to male dominance as size increased. No significant relationship between length and batch fecundity was found. The population has a spawning period from February to August. Overall, 54.4% of macroscopic evaluations were incorrect compared to histological results. Of the errors, 14.8% were a failure to detect ovotestes, 12.7% were classifying non-gonadal tissue as ovary or testis, 12.2% were misclassifying sex, and 12.7% were misclassifying maturity status. However, the largest source of error (47.7%) was from misclassifying both sex and maturity status. Of these, 92.9% were macroscopically classified as immature females, but were histologically confirmed to be mature males. Compared to histological results, the only accurate macroscopic results were the absence of a sex-based difference in weight-length relationship and spawning seasonality estimated by a gonadosomatic index (February-June). The use of macroscopic methods to estimate reproductive life-history parameters for sex-changing reefes fish may introduce significant inaccuracies and misinterpretations. Of the parameters estimated by histological methods, size at maturity, size-specific sex ratios, and spawning seasonality have the greatest potential to inform local fishery management policy.

Movement patterns of a small-bodied minnow suggest nomadism in a fragmented, desert river.

BACKGROUND: Unfettered movement among habitats is crucial for fish to access patchily distributed resources and complete their life cycle, but many riverscapes in the American Southwest are fragmented by dams and dewatering. The endangered Rio Grande silvery minnow (Hybognathus amarus, RGSM) persists in a fragmented remnant of its former range (ca. 5%), and its movement ecology is understudied. METHODS: We tracked movements of hatchery-reared RGSM, tagged with passive integrated transponder tags, using stationary and mobile antennas from 2019 to 2022. We quantified probability of movement and total distance moved by RGSM released above and below a dam. We then assessed how well two prevailing riverine movement theories (i.e., restricted movement paradigm [RMP] and colonization cycle hypothesis [CCH]) explained RGSM movement patterns. RESULTS: We detected 36.8% of released RGSM (n = 37,215) making at least one movement. Movements were leptokurtic and substantially greater than expected based on the RMP for both stationary (1.7-5.9 m) and mobile (30.3-77.8 m) individuals. On average, RGSM were detected at large for 75 days and moved a total of 12.2 rkm within a year. The maximum total distance moved by RGSM was 103 rkm. Similarly, we observed a multimodal distribution of detected range sizes with a mean detected range of 2.4 rkm and a maximum detected range of 78.2 rkm. We found little support for an upstream movement bias, as expected under the CCH, and most movements (74%) were directed downstream. CONCLUSIONS: Our data suggest RGSM are highly mobile, with the ability to make long-distance movements. Neither movement theory adequately described movement patterns of RGSM; instead, our findings support a nomadic movement pattern and an apparent drift paradox matching recent studies of other pelagic-broadcast spawning minnows where populations persist upstream despite experiencing downstream drift as larvae. Resolution of the drift paradox may be achieved through further, targeted studies into different aspects of the species' life history. Quantification of RGSM movement provides crucial insights into the species' movement ecology and may help define the appropriate scale of recovery efforts.

Long-term aquarium records delineate the synchronized spawning strategy of Acropora corals.

Aligning spawning timing with seasonal environmental changes is critical for both terrestrial and aquatic organisms. However, mechanisms to regulate reproductive activity in response to environmental factors are not well understood, partly owing to the technical difficulty in maintaining detailed long-term observational data of the reproductive activities in the same population across years. In this study, we present an application of the aquarium system to examine the long-term spawning properties of corals. Spawning records over a 15-year period at the Okinawa Churaumi Aquarium revealed that the spawning timing of Acropora corals in aquarium tanks aligned well with that of wild corals from a neighbouring reef. Using the aquarium population as a model, we investigated the relationship between key environmental factors and the timing of the first and peak spawning dates of Acropora corals during a 15-year period. The results suggest that the spawning window of each spawning season is largely influenced by the water temperature and that the timing of peak spawning can be fine-tuned in response to environmental fluctuations. This behavioural feature can prevent synchronous spawning events during unfavourable environmental conditions and increase long-term reproductive reliability.

Histological characteristics of oocyte differentiation in the captive longnose seahorse Hippocampus trimaculatus (Leach, 1814).

This study aimed to explore the reproductive histology and oocyte differentiation of the longnose seahorse Hippocampus trimaculatus (Leach, 1814) in captivity. Five mature healthy females were histologically observed. The reproductive systems of the five specimens exhibited similar morphological characteristics with a pair of saccular creamy white ovaries merging caudally into a single gonoduct. There were two germinal ridges lined with a layer of germinal epithelium (GE). The ovarian maturation of this species was considered asynchronous. The oogenic cells were classified into oogonia and oocytes at several developmental phases based on their size and characteristics. Oogonia were identified among the connective tissue in the middle area of the GE. The stromal compartment contained oocytes that were classified into four distinct phases: the primary growth (PG) phase having two steps (perinucleolar and oil droplets-cortical alveolar steps) and the secondary growth (SG) phase with three oocyte types, including early SG oocytes, late SG oocytes, and fully grown oocytes. The atretic oocytes (AO) were observed in all stages of oogenesis. Postovulatory follicles were also seen among the ovarian connective tissue. The occurrence of postovulatory follicles suggested that the specimens analysed in this study were in the spawning period. This research provides new insights into the identification of the reproductive cycles and morphological characteristics of the ovary of H. trimaculatus.

Seasonal migrations of the European sea bass (Dicentrarchus labrax L.) in UK and surrounding waters.

The movements and behaviour of mature European sea bass (Dicentrarchus labrax L.) in UK waters have not been studied extensively since a series of mark-recapture experiments during the 1970s, 80s and 90s. To better understand the timing and extent of seasonal migrations, 171 mature sea bass > 42 cm were internally tagged with floated electronic tags programmed to record temperature and depth, and released in the English Channel, in the southern North Sea and in the Irish Sea. Among the 48 tags returned to date, sea bass were at liberty for 370 ± 337 days and were recovered 172 ± 200 km from their respective release locations. Most tags were recovered from beaches (54%), or via the fishery (44%). A comparison of the reconstructed tracks from returned electronic tags with the recapture locations of 237 mark-recapture returns (6.5%) from 3615 sea bass released between 1970 and 2020 showed strong overlap. Seasonal movements between shallow areas (Q2-Q3) and deeper spawning areas (Q4-Q1) were accompanied by elevated vertical swimming speeds and average water temperatures of 8.5 °C in the English Channel and Irish Sea, but lower temperatures in the North Sea. Movements between the Celtic Sea/Irish Sea and the North Sea and vice versa demonstrate high levels of connectivity in UK waters. We demonstrate that a proportion of sea bass remained resident within the North Sea throughout the year, with a strong suggestion that spawning might be occurring. These data have significant implications for the future sustainable management of sea bass stocks in UK and surrounding waters.

Early arrival of spring-spawning Atlantic herring Clupea harengus at their spawning ground in the Kiel Fjord, western Baltic, relates to increasing winter seawater temperature.

The disturbance of marine organism phenology due to climate change and the subsequent effects on recruitment success are still poorly understood, especially in migratory fish species, such as the Atlantic herring (Clupea harengus; Clupeidae). Here we used the commercial catch data from a local fisher over a 50-year period (1971-2020) to estimate western Baltic spring-spawning (WBSS) herring mean arrival time Q50 (i.e., the week when 50% of the total fish catches had been made) at their spawning ground within the Kiel Fjord, southwest Baltic Sea, and the duration of the spawning season for each year. The relationship between the seawater temperature in the Kiel Bight and other environmental parameters (such as water salinity, North Atlantic and Atlantic multidecadal oscillations) and Q50 was evaluated using a general linear model to test the hypothesis that fish arrived earlier after warm than cold winters. We also estimated the accumulated thermal time to Q50 during gonadal development to estimate the effects of seawater temperature on the variations of Q50. The results of this study revealed a dramatic decrease in herring catches within the Kiel Fjord since the mid-1990s, as documented for the whole southwestern Baltic Sea. Warmer winter seawater temperature was the only factor related to an earlier arrival (1 week for one January seawater temperature degree increase) of herring at their spawning ground. The relationship was found for the first time on week 52 of the year prior to spawning and was the strongest (50% of the variability explained) from the fourth week of January (8 weeks before the mean Q50 among the studied years). A thermal constant to Q50 (~316°C day) was found when temperatures were integrated from the 49th week of the year prior to spawning. These results indicate that seawater temperature enhanced the speed of gonadal maturation during the latest phases of gametogenesis, leading to an early fish arrival under warm conditions. The duration of the spawning season was elongated during warmer years, therefore potentially mitigating the effects of trophic mismatch when fish spawn early. The results of this study highlight the altering effects of climate change on the spawning activity of a migratory fish species in the Baltic Sea where fast global changes presage that in other coastal areas worldwide.

Effect of hormonal treatment on artificial propagation, spawning performance and embryonic development of striped snakehead Channa striata (Bloch, 1793).

Present study describes the spawning induction of striped Snakehead, Channa striata using carp pituitary extract (CPE) and LH-RH agonist i.e. Buserelin (Glp-His-Trp-Ser-Tyr-Ser-tBu-Leu-Arg-Pro-NHEt). Total four treatments were designed under both hormones trail and treated as control group, T1, T2, and T3 with three replications of each treatment. While breeders under all hormone treatments showed spawning performances, no spawning performance was observed in control group. Latency time after hormonal treatment was lowest (20-24 hrs) in case of CPE than Buserelin (25-30 hrs). Regarding to CPE, spawning, fertilization and hatching rate were higher with the increasing doses of CPE in different treatments. The highest mean ± Standard deviation spawning, fertilization and hatching rate were 85.60±8.58 %, 79.38±4.89 % and 64.33±6.60 % respectively in T3 where dose of CPE was 80 mg kg-1. Similarly, in case of Buserelin hormone highest spawning rate was found in T3 (80.61±5.59) where dose of Buserelin was 0.80 µg kg-1 body weight. Fertilization rate was on the level 48.57±5.99, 70.62±5.33 and 90.32±4.79 respectively for T1, T2, and T3.Whilst, hatching rate was found 20.81±4.91, 37.11±4.50 and 61.33±6.61 in T1, T2, and T3 treatments respectively. However, T3 exhibited best performance regarding spawning, fertilization and hatching rate which were significantly higher than other two treatments.The current study revealed that spawning induction using carp pituitary extract and Buserelin is effective and might be useful for artificial breeding of Channa striata. Regarding to dose application i.e. 80 mg kg-1 of CPE and 0.80 µg kg-1 of Buserelin may be successfully applied to ovulation stimulation of Channa striata.

Recruitment of European sea bass (Dicentrarchus labrax) in northerly UK estuaries indicates a mismatch between spawning and fisheries closure periods.

European sea bass (Dicentrarchus labrax) is a species of high commercial and recreational value, but it exhibits highly variable recruitment rates and has been subject to recent declines. Emergency management measures put in place to protect spawning stocks include the annual closure of commercial and recreational fisheries over a 2-month, February-March, window. Whether this protection measure is having the desired outcome for this data-poor species remains unclear. Otolith microstructural analyses (counts and widths of daily growth rings and check marks indicative of settlement) were used to estimate (1) spawn timing, (2) pelagic larval duration and settlement timing, (3) growth rate and condition, and (4) the otolith-fish size relationship for juvenile European sea bass caught from two estuaries in Wales (Dwyryd, Y Foryd), located at the northern edge of the species range. We observed a significant mismatch between the timing of fisheries closures and the spawning, with 99.2% of recruits having been spawned after the fishery had reopened (back-calculated median spawn date = May 5 ± 17 days SD), suggesting that the closure may be too early to adequately protect this population. Further, we present the first empirically derived estimates of pelagic larval duration for sea bass recruits settling in UK habitats, which showed a strong negative relationship with spawn date. Finally, we found significant differences in fish condition between the two estuaries, suggesting local variation in habitat quality. The results suggest that the timing of current fisheries closures may not be adequately protecting the spawners supplying these northernmost estuaries, which are likely to become increasingly important as sea bass distributions shift northward in our climate future.

Quantitative determination of flow rate variations in reservoir Eco-scheduling: A case study of Yangqu dam in the upper yellow river.

Accurate quantification of flow dynamics during reservoir ecological scheduling hinders the maintenance of normal reproductive activities in downstream riverine fish. This study proposed a quantitative method for determining the flow rate changes in reservoir ecological scheduling. The approach utilized the daily flow rate and daily flow-rate increment to characterize the flow process. Adopting the perspective of shifting spawning grounds of adhesive egg-laying fish species in response to flow rate variations, we introduced the Spawning Ground Overlap Rate as an indicator and utilized it to determine flow rate changes. Focusing on the downstream area of the Yangqu Hydropower Station in the upper reaches of the Yellow River, we calculated the distribution of spawning grounds and the Spawning Ground Overlap Rate in the region. We set a threshold for the Spawning Ground Overlap Rate to restrict the flow rate changes. The results indicated that during the fish spawning period, the ecological flow range in the downstream area of the Yangqu Dam was 480-1200 m3/s. It was required to maintain a daily flow rate change of less than 49.45 m3/(s·d) and a maximum seven-day flow difference of less than 227.76 m3/s to maintain the optimal level of spawning ground overlap rate. Additionally, it was necessary to keep the daily flow rate change below 123.83 m3/(s·d) and the maximum seven-day flow difference below 368.84 m3/s to maintain the minimum spawning ground overlap rate. The findings provide foundational data for determining flow dynamics during the ecological scheduling of the spawning period for viscous-spawning fish.

A rare case of intra-ovarian oocyte maturation.

The intra-ovarian presence of ootids, i.e. female gametes that have completed meiosis, is considered exceptional in the animal kingdom. The present study explores the first such case to be reported in a sea cucumber (Echinodermata: Holothuroidea). In the overwhelming majority of animals, including holothuroids, oocytes (i.e. immature female gametes) that are developing in the ovary undergo a primary arrest at the prophase stage of meiosis, which may last from days to decades. In free-spawning taxa, this arrest is normally lifted only during or shortly before transit in the gonoduct, when gamete release (spawning) is imminent. However, oocytes of the holothuroid Chiridota laevis were discovered to have resumed the second meiotic division including the completion of germinal vesicle breakdown and polar-body expulsion inside the ovary, effectively reaching the ootid stage concomitantly with ovulation (i.e. escape from follicle cells) prior to spawning. The potential drivers and significance of this exceptionally rare case of full intra-ovarian oogenic maturation are discussed.

Temporal declines in fecundity: A study of southern Gulf of St. Lawrence Atlantic herring (Clupea harengus) and implications for potential reproductive output.

Individuals must reproduce to survive and thrive from generation to generation. In fish, the fecundity of individuals and estimates of total reproductive output are critical for evaluating reproductive success and understanding population dynamics. Estimating fecundity is an onerous task; therefore, many populations lack contemporary estimates of fecundity and size-fecundity relationships. However, reproductive dynamics are not static in time; therefore, it is important to develop contemporary fecundity estimates to better inform conservation and management action. To highlight the importance of contemporary fecundity estimates, we examined the fecundity of southern Gulf of St. Lawrence (sGSL) spring and fall spawning Atlantic herring in 2022, developed size-fecundity models, and compared these to historical fecundity estimates and models. Our results suggest that the average fecundity of sGSL spring and fall herring has undergone a substantial temporal decline of approximately 47% and 58%, respectively, since the 1970s and 1980s. The size-fecundity relationships for fall spawning herring have shifted, with fish of a given size exhibiting lower fecundity in 2022 compared to the 1970s. Alternatively, the size-fecundity relationships for spring spawning herring have remained relatively static. Furthermore, simulations highlighted a substantial reduction in potential reproductive output in 2022 compared to 1970 of approximately 32% and 68% for spring and fall spawners, respectively, based on a fixed number of mature females, which may have negative implications for stock rebuilding. Overall, our study provides support for periodic estimates of fecundity in fish populations to better understand temporal changes in reproductive and population dynamics.

Influence of temperature on hatching success of skipjack tuna (Katsuwonus pelamis): Implications for spawning availability of warm habitats.

Exploring the effects of water temperature on egg development in skipjack tuna (Katsuwonus pelamis) has substantial implications for evaluations of wild spawning habitats. In the present study, we examined the hatching success and duration as a function of temperature from 21 to 33°C under captive environments. A high hatching rate of over 50% between 23 and 31°C was observed, with the shortest hatching duration at 31°C. Because the egg period is vulnerable to predators, a shortened hatching duration with warming water would be ecologically advantageous for K. pelamis, as its main spawning grounds are located in tropical areas.

Exploring the reproductive features of Clupisoma garua from Ganges River, Bangladesh: insights into eco-climatic factors for sustainable management and advancements in aquaculture practices.

The Garua bacha, Clupisoma garua, holds considerable commercial and nutritional significance as a fish species. This study investigates the relationships between GSI (gonadosomatic index) and eco-climatic conditions, and provides comprehensive insights into several facets of reproduction, such as size at first sexual maturity (Lm), spawning season, peak spawning period, and proposes sustainable management strategies for C. garua in the Ganges River, northwestern, Bangladesh. Since January to December 2017, 570 female individuals have been gathered from the Ganges River using cast nets, gill nets, and square lift nets on a monthly basis. The total length (TL) of the specimens varied from 3.90 to 26.30 cm. Lm was estimated as 14.30 cm based on the results of the TL vs. GSI, MGSI (modified gonadosomatic index), DI (Dobriyal index), 14.00 cm through TL-FL regressions, and 15.18 cm depending on Lmax for this species. Furthermore, the spawning period was observed from May to August, peaking in June and July, based on greater GSI, MGSI, and DI values. Throughout the peak spawning season, there was not a significant differ from a value of 100 in the relative weight (WR) of the female species. During the spawning season, a substantial association between temperature and GSI (p = 0.0038); rainfall and GSI (p = 0.0043); DO and GSI (p = 0.0043); pH and GSI (p = 0.0002); and alkalinity and GSI (p = 0.0001) was detected. Analyzing a 55-year data series (1964-2018), it became clear that an increase in average air temperature of 0.0289 °C and a decrease in rainfall of 2.988 mm per year might possibly delay the spawning season of this species. As a consequence, the results of this study provide crucial information for developing management strategies to safeguard C. garua populations in the Ganges River and its adjacent ecosystems.

Lunar Age-Dependent Oscillations in Expression of the Genes for Kisspeptin, GnIH, and Their Receptors in the Grass Puffer during the Spawning Season.

Grass puffer is a semilunar-synchronized spawner: spawning occurs on beaches only for several days of spring tide around new moon (lunar age 0) and full moon (lunar age 15) every 2 weeks from spring to early summer. To investigate the role of kisspeptin and gonadotropin-inhibitory hormone (GnIH) in the semilunar-synchronized spawning, lunar age-dependent expression of the genes encoding kisspeptin (kiss2), kisspeptin receptor (kissr2), GnIH (gnih), GnIH receptor (gnihr), gonadotropin-releasing hormone 1 (GnRH1) (gnrh1), and three gonadotropin (GTH) subunits (gpa, fshb, lhb) was examined in the male grass puffer, which was kept in an aquarium under natural light condition in a lunar month during the spawning period. In the brain, both kiss2 and kissr2 showed lunar variations with a peak at lunar age 10, while both gnih and gnihr showed semilunar variations with two peaks at lunar age 0 and 20. On the other hand, gnrh1 showed semilunar variation with two peaks at lunar age 0 and 15. In the pituitary, kiss2, kissr2, gnih, and gnihr showed similar variations to those shown in the brain. The fshb and lhb mRNA levels showed semilunar variations with two peaks at lunar age 0 and 15. The present study shows lunar and semilunar oscillations of kiss2/kissr2 and gnih/gnihr expressions, respectively, with their peaks around spring tide in the brain and pituitary along with the semilunar expressions of gnrh1 and the pituitary GTH subunit genes. These results suggest that the lunar age-dependent expressions of the kisspeptin, GnIH, and their receptor genes may be primarily important in the control of the precisely timed semilunar spawning of the grass puffer.

Gene expression profiles of Japanese precious coral Corallium japonicum during gametogenesis.

Background: Corallium japonicum, a prized resource in Japan, plays a vital role in traditional arts and fishing industries. Because of diminished stock due to overexploitation, ongoing efforts are focused on restoration through transplantation. This study aimed to enhance our understanding of the reproductive biology of these valuable corals and find more efficient methods for sex determination, which may significantly contribute to conservation initiatives. Methods: We used 12 three-month aquarium reared C. japonicum colony fragments, conducted histological analysis for maturity and sex verification, and performed transcriptome analysis via de novo assembly and mapping using the C. rubrum transcriptome to explore gene expression differences between female and male C. japonicum. Results: Our histological observations enabled sex identification in 33% of incompletely mature samples. However, the sex of the remaining 67% of samples, classified as immature, could not be identified. RNA-seq yielded approximately 21-31 million short reads from 12 samples. De novo assembly yielded 404,439 highly expressed transcripts. Among them, 855 showed significant differential expression, with 786 differentially expressed transcripts between females and males. Heatmap analysis highlighted 283 female-specific and 525 male-specific upregulated transcripts. Transcriptome assembly mapped to C. rubrum yielded 28,092 contigs, leading to the identification of 190 highly differentially expressed genes, with 113 upregulated exclusively in females and 70 upregulated exclusively in males. Blastp analysis provided putative protein annotations for 83 female and 72 male transcripts. Annotation analysis revealed that female biological processes were related to oocyte proliferation and reproduction, whereas those in males were associated with cell adhesion. Discussion: Transcriptome analysis revealed sex-specific gene upregulation in incompletely mature C. japonicum and shared transcripts with C. rubrum, providing insight into its gene expression patterns. This study highlights the importance of using both de novo and reference-based assembly methods. Functional enrichment analysis showed that females exhibited enrichment in cell proliferation and reproduction pathways, while males exhibited enrichment in cell adhesion pathways. To the best of our knowledge, this is the first report on the gene expressions of each sex during the spawning season. Our findings offer valuable insights into the physiological ecology of incompletely mature red Japanese precious corals and suggest a method for identifying sex using various genes expressed in female and male individuals. In the future, techniques such as transplantation, artificial fertilization, and larval rearing may involve sex determination methods based on differences in gene expression to help conserve precious coral resources and ecosystems.