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Surrogate broodstock technology facilitates the production of donor-derived gametes in surrogates, and comprises transplanting germ cells of a donor into recipients of a different strain or different species. The following applications of this technology are expected in the field of aquaculture: (1) the efficient and reliable production of offspring carrying superior genetic traits by transplanting donor germ cells from a single selected fish with superior traits into many recipient fish; (2) the reduction of the time required to breed fish by using a recipient species with a short generation time to produce gametes of a species with a long generation time; (3) the long-term storage of valuable species or strains as genetic resources by cryopreserving germ cells for transplantation; (4) the mass production of genetically sterile fish by transplanting germ cells of a donor fish that is sterile due to a mutation in the somatic cells into normal recipients without this mutation. It is expected that a combination of these techniques will greatly accelerate the breeding of aquaculture species. It is important to adapt surrogate broodstock technology to a wider range of fishery species and further improve the efficiency of donor-derived gamete production when using surrogate broodstock.

Bitterling is a small cyprinid fish facing an increasing risk of extinction owing to habitat destruction and decreasing freshwater mussel population that are used as their spawning substrates. Owing to their large size and high yolk contents, methods for cryopreservation of their eggs or embryos, which is a promising method for long-term preservation of their genetic resources, are still not available. We conducted this study to evaluate the feasibility of gamete production by transplanting cryopreserved testicular cells into germ cell–less recipients that were produced by knockdown of dead end gene. Immature testes isolated from recessive albino Chinese rosy bitterlings were cryopreserved by slow freezing or vitrification. Approximately 3000 slow-frozen or vitrified cells were transplanted into the peritoneal cavity of 4-day-old germ cell–less wild-type Chinese rosy bitterlings. We observed no significant differences in the incorporation rates of the slow-frozen and vitrified cells into the genital ridges of recipients compared with those of freshly prepared cells. When the recipients matured, almost half of the male or female recipients that received freshly prepared, slow-frozen, or vitrified cells produced gametes derived from donor cells, with no significant differences in their fecundity among the 3 groups. Moreover, fertilization of the resulting eggs and sperm produced donor-derived offspring exhibiting the albino phenotype. Therefore, the abovementioned methods could be used as a powerful and practical method for long-term preservation of bitterling genetic resources for biotic conservation.

Many bitterling species are facing extinction because of habitat destruction. Since cryopreservation of fish eggs is still not available to date due to their large size and high yolk content, long-term and stable storage of bitterling genetic resources is currently not possible. We recently discovered that cryopreservation of early-stage germ cells is possible in several fish species and that functional gametes derived from the frozen materials can be produced through their transplantation to embryonic recipients. However, bitterlings have uniquely shaped eggs and their embryos are extremely fragile, making it difficult to perform germ cell transplantation. Therefore, as a first step, we conducted intra-species spermatogonial transplantation using recessive albino Chinese rosy bitterling as donors and wild-type Chinese rosy bitterling as recipients to develop a system to convert freezable early-stage germ cells into functional gametes, particularly eggs. Approximately 3000 testicular cells were transplanted into the peritoneal cavity of 4-day-old germ cell-less recipient embryos produced by dead end (dnd)-knockdown. At 6 months, ten male recipients and nine female recipients produced gametes. Mating studies with the opposite sex of recessive albino control fish revealed that six males and three females produced only albino offspring, suggesting that these recipients' endogenous germ cells were completely removed by dnd-knockdown and they produced only donor-derived gametes. Thus, we successfully established a germ cell transplantation system in an iconic endangered teleost, bitterling. The technology established in this study can be directly applied to produce functional gametes of endangered bitterlings using cryopreserved donor cells. Summary Sentence Allogeneic germ cell transplantation system was established using germ cell-less embryonic recipients produced by dead end-knockdown in a bitterling, which is an iconic endangered fish species.

Stem cell maintenance is established by neighboring niche cells that promote stem cell self-renewal. However, it is poorly understood how stem cell activity is regulated by systemic, tissue-extrinsic signals in response to environmental cues and changes in physiological status. Here, we show that neuropeptide F (NPF) signaling plays an important role in the pathway regulating mating-induced germline stem cell (GSC) proliferation in the fruit fly Drosophila melanogaster. NPF expressed in enteroendocrine cells (EECs) of the midgut is released in response to the seminal-fluid protein sex peptide (SP) upon mating. This midgut-derived NPF controls mating-induced GSC proliferation via ovarian NPF receptor (NPFR) activity, which modulates bone morphogenetic protein (BMP) signaling levels in GSCs. Our study provides a molecular mechanism that describes how a gut-derived systemic factor couples stem cell behavior to physiological status, such as mating, through interorgan communication.

The conservation of endangered fish is of critical importance. Cryobanking could provide an effective backup measure for use in conjunction with the conservation of natural populations; however, methodology for cryopreservation of fish eggs and embryos has not yet been developed. The present study established a methodology capable of deriving functional eggs and sperm from frozen type A spermatogonia (ASGs). Whole testes taken from rainbow trout were slowly frozen in a cryomedium, and the viability of ASGs within these testes did not decrease over a 728-d freezing period. Frozenthawed ASGs that were intraperitoneally transplanted into sterile triploid hatchlings migrated toward, and were incorporated into, recipient genital ridges. Transplantability of ASGs did not decrease after as much as 939 d of cryopreservation. Nearly half of triploid recipients produced functional eggs or sperm derived from the frozen ASGs and displayed high fecundity. Fertilization of resultant gametes resulted in the successful production of normal, frozen ASG-derived offspring. Feasibility and simplicity of this methodology will call for an immediate application for real conservation of endangered wild salmonids.

Many freshwater fish face extinctions due to habitat destruction and overfishing. Bitterlings, which lay their eggs in the gill chambers of freshwater mussels, are in steep decline worldwide, along with mussels, owing to the concreting and siltation of riverbeds. In particular, Tokyo bitterling is listed as “endangered” on the Red List and is on the verge of extinction. Although in situ and ex situ conservation efforts are underway, their effectiveness is limited. Here, we developed a method for long-term preservation of genetic resources by combining germ cell cryopreservation and transplantation. We transplanted Tokyo bitterling germ cells stored in liquid nitrogen into oily bitterling recipients lacking endogenous germ cells due to dnd knockdown, and the recipients matured and repeatedly produced many eggs and sperm. The resulting offspring had the Tokyo bitterling genotype and morphology. This study shows that even extinct species can be revived if their germ cell bank is available.

Invasive alien fish constitute a major problem in the conservation of aquatic ecosystems. The most common method of exterminating invasive fish is physical capture by fishing gear; however, it is extremely difficult to eradicate them. Here, we developed a novel methodology to inhibit the reproduction of invasive fish by releasing surrogate males of the same species, which produce sperm of different species, into rivers, allowing them to produce lethal hybrids through natural spawning. This study focused on rainbow trout, an invasive fish that negatively impacts ecosystems worldwide. Since rainbow trout × brown trout hybrids are lethal, surrogate rainbow trout producing brown trout sperm were created by germline stem cell transplantation. Releasing the surrogate males into an experimental river inhabited by wild-type rainbow trout females revealed that they naturally mated and produced lethal hybrid offspring. This is a novel and powerful methodology for eradicating invasive fish.

Colonization of new ecological niches has triggered large adaptive radiations. Although some lineages have made use of such opportunities, not all do so.The factors causing this variation among lineages are largely unknown. Here, we show that deficiency in docosahexaenoic acid (DHA), an essential ω-3 fatty acid, can constrain freshwater colonization by marine fishes. Our genomic analyses revealed multiple independent duplications of the fatty acid desaturase gene Fads2 in stickleback lineages that subsequently colonized and radiated in freshwater habitats, but not in close relatives that failed to colonize.Transgenic manipulation of Fads2 in marine stickleback increased their ability to synthesize DHA and survive on DHA-deficient diets. Multiple freshwater ray-finned fishes also show a convergent increase in Fads2 copies, indicating its key role in freshwater colonization.

There is great variation in the size and shape of teleost eggs from species to species. The size of the teleost egg depends on the amount of yolk accumulated in the egg, which is an important factor directly affecting the survival of hatchlings. Egg shape also contributes significantly to spawning ecology and survival during the prehatching stage. In this study, we used bitterlings, which show a wide variety of egg volumes and shapes, to elucidate whether these factors are determined by germ cells or somatic cells. Reciprocal transplantations of germ cells between two bitterling species revealed that the egg volume was identical to that of the germ cell donor species in both combinations. The egg shape was also very similar to that of the species providing the germ cells. These results suggest that the volume and shape of teleost eggs are greatly influenced by germ cell autonomy.

Pacific bluefin tuna ( Thunnus orientalis ) remains heavily depleted due to overexploitation. Aquaculture and stock enhancement based on artificial seedlings could be effective solutions to this problem. However, widespread adoption of seedling production is limited because spawning in captivity of bluefin tuna, a large pelagic top predator, requires much space, time, cost, and labour. To downsize and shorten their generation time, here, we show that xenogeneic germ cell transplantation enables a surrogate production of functional bluefin tuna sperm in hybrid little tuna (genus: Euthynnus ), which attain sexual maturity at a smaller body size and shorter time. Intraperitonially transplanted bluefin tuna germ cells migrate towards and are incorporated into the gonads of little tuna larvae. These recipients produce functional donor-derived bluefin tuna sperm at a mere 8 months of age and 1 kg of body size. This result represents an advance toward compact, low-cost, and time-efficient seedling production that will improve the efficiency of bluefin tuna aquaculture. Here, the authors show that germ cell transplantation and surrogate production of bluefin tuna sperm in small-bodied fish species shortens the period in which sperm is produced. This study opens the possibility of highly efficient seed production in bluefin tuna.