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Interspecific hybridization may trigger the transition from sexual reproduction to asexuality, but mechanistic reasons for such a change in a hybrid’s reproduction are poorly understood. Gametogenesis of many asexual hybrids involves a stage of premeiotic endoreplication (PMER), when gonial cells duplicate chromosomes and subsequent meiotic divisions involve bivalents between identical copies, leading to production of clonal gametes. Here, we investigated the triggers of PMER and whether its induction is linked to intrinsic stimuli within a hybrid’s gonial cells or whether it is regulated by the surrounding gonadal tissue. We investigated gametogenesis in the Cobitis taenia hybrid complex, which involves sexually reproducing species (Cobitis elongatoides and C. taenia) as well as their hybrids, where females reproduce clonally via PMER while males are sterile. We transplanted spermatogonial stem cells (SSCs) from C. elongatoides and triploid hybrid males into embryos of sexual species and of asexual hybrid females, respectively, and observed their development in an allospecific gonadal environment. Sexual SSCs underwent regular meiosis and produced normally reduced gametes when transplanted into clonal females. On the other hand, the hybrid’s SSCs lead to sterility when transplanted into sexual males but maintained their ability to undergo asexual development (PMER) and production of clonal eggs, when transplanted into sexual females. This suggests that asexual gametogenesis is under complex control when somatic gonadal tissue indirectly affects the execution of asexual development by determining the sexual differentiation of stem cells and once such cells develop to female phenotypes, hybrid germ cells trigger the PMER from their intrinsic signals.

In electroreceptive jawed fishes and amphibians, individual lateral line placodes form lines of neuromasts on the head containing mechanosensory hair cells, flanked by fields of ampullary organs containing electroreceptors - modified hair cells that respond to weak electric fields. Extensively shared gene expression suggests that conserved molecular mechanisms are involved in their development, but a few transcription factor genes are restricted either to the developing electrosensory or mechanosensory lateral line. Here, we used CRISPR/Cas9-mediated mutagenesis in F0-injected sterlet embryos (Acipenser ruthenus, a sturgeon) to test the function of three such genes. We found that the ‘hair cell’ transcription factor geneAtoh1is required for both hair cell and electroreceptor differentiation in sterlet, and forPou4f3andGfi1expression in both neuromasts and ampullary organs. These data support the conservation of developmental mechanisms between hair cells and electroreceptors. Targeting ampullary organ-restrictedNeurod4did not yield any phenotype, potentially owing to redundancy with otherNeurodgenes that we found to be expressed in sterlet ampullary organs. After targeting mechanosensory-restrictedFoxg1, ampullary organs formed within neuromast lines, suggesting that Foxg1 normally represses their development. We speculate that electrosensory organs may be the ‘default’ fate of lateral line primordia in electroreceptive vertebrates.

Common carp (

The aim of this study was to test whether vitrification of sterlet Acipenser ruthenus and Russian sturgeon Acipenser gueldenstaedtii ovarian tissue through needle-immersed vitrification (NIV) is an efficient strategy for the preservation of oogonia (OOG) in order to supplement the current conservation efforts for these endangered fish species. Histological analyses of the gonads displayed that the ovaries of both species were immature and contained predominantly OOG and primary oocytes. The germline origin of these cells was verified by localization of the vasa protein through immunocytochemistry. NIV protocol was optimized by testing different equilibration (ES) and vitrification solutions (VS) containing various concentrations of dimethyl sulfoxide (Me

The cleavage pattern of a vertebrate’s embryo is either holoblastic (complete) or meroblastic (partial). Sturgeon and other basal bony fishes represent a transition of the cleavage pattern. To understand the transition, it is essential to develop an effective technique for the inhibition of specific blastomere cleavage. So far, various studies have demonstrated that diatom-derived polyunsaturated aldehyde (PUA), 2,4-decadienal (DD)—a model aldehyde for experimental studies—adversely affects the developing embryos of several aquatic species. In this study, we employed DD for inhibition of cleavage of a definite blastomere in sturgeon embryos under various conditions. The effective treatment was found to be a combination of DD injection (0.01 v/v) and visible light (44.86–91.15 W m−2). Notably, DD injection or light irradiation alone cannot inhibit cleavage. Furthermore, spatial RNA localization analysis using quantitative polymerase chain reaction (qPCR)-tomography revealed that the localized pattern of selected maternal messenger ribonucleic acids (mRNAs) remained constant along the animal–vegetal (A-V) axis, which suggests that RNA localization is completed by the end of oogenesis and that early embryonic cleavage is not required for A-V asymmetry preservation.

The zebrafish (Danio rerio) genome contains a single gene fads2 encoding a desaturase (FADS2) with both Δ6 and Δ5 activities, the key player in the endogenous biosynthesis of long-chain polyunsaturated fatty acids (LC-PUFAs), which serve essential functions as membrane components, sources of energy and signaling molecules. LC-PUFAs include the precursors of eicosanoids and are thus predicted to be indispensable molecules for reproductive health in virtually all vertebrates. In mice, an amniotic vertebrate, fads2 deletion mutants, both males and females, have been confirmed to be sterile. In anamniotic vertebrates, such as fish, there is still no information available on the reproductive (in)ability of fads2 mutants, although zebrafish have become an increasingly important model of lipid metabolism, including some aspects of the generation of germ cells and early embryonic development. In the present study, we apply the CRISPR/Cas9 genome editing system to induce mutations in the zebrafish genome and create crispants displaying a degree of fads2 gene editing within the range of 50–80%. Focusing on adult G0 crispant females, we investigated the LC-PUFA profiles of eggs. Our data suggest an impaired pathway of the LC-PUFA biosynthesis of the ω6 and ω3 series in the first-rate limiting steps of the conversion of linoleic acid (LA) into γ-linolenic acid (GLA), and α-linolenic acid (ALA) into stearidonic acid (SDA), respectively, finally resulting in bad-quality eggs. Our data suggest the existence of an alternative Δ8 pathway, which bypasses the first endogenous LC-PUFA biosynthetic step in zebrafish in vivo, and suggest that the zebrafish bifunctional FADS2 enzyme is actually a trifunctional Δ6/Δ5/Δ8 desaturase.

DNA damage during early life stages may have a negative effect on embryo development, inducing mortality and malformations that have long-lasting effects during adult life. Therefore, in the current study, we analyzed the effect of DNA damage induced by genotoxicants (camptothecin (CPT) and olaparib) at different stages of embryo development. The survival, DNA fragmentation, transcriptome, and proteome of the endangered sturgeon

The zebrafish (

Interspecific hybridization may trigger the transition from sexual reproduction to asexuality, but mechanistic reasons for such a change in a hybrid’s reproduction are poorly understood. Gametogenesis of many asexual hybrids involves a stage of premeiotic endoreduplication (PMER), when gonial cells duplicate chromosomes and subsequent meiotic divisions involve bivalents between identical copies, leading to production of clonal gametes. Here, we investigated the triggers of PMER and whether its induction is linked to intrinsic stimuli within a hybrid’s gonial cells or whether it is regulated by the surrounding gonadal tissue.We investigated gametogenesis in theCobitis taeniahybrid complex, which involves sexually reproducing species (Cobitis elongatoidesandC. taenia) as well as their hybrids, where females reproduce clonallyviaPMER while males are sterile. We transplanted spermatogonial stem cells (SSCs) fromC. elongatoidesand triploid hybrid males into embryos of sexual species and of asexual hybrid females, respectively, and observed their development in an allospecific gonadal environment. Sexual SSCs underwent regular meiosis and produced normally reduced gametes when transplanted into clonal females. On the other hand, the hybrid’s SSCs lead to sterility when transplanted into sexual males, but maintained their ability to undergo asexual development (PMER) and production of clonal eggs, when transplanted into sexual females.This suggests that asexual gametogenesis is under complex control when somatic gonadal tissue indirectly affects the execution of asexual development by determining the sexual differentiation of stem cells and once such cells develop to female phenotypes, hybrid germ cells trigger the PMER from their intrinsic signals.Significance StatementAlthough sexual reproduction is a dominant trait among all eukaryotes, many taxa have evolved the ability to reproduce asexually. While asexuality appears to be linked to interspecific hybridization, it remains unknown how the coexistence of diverged genomes may initiate such a swap in reproduction. In our study, we transplanted germ cells between asexual hybrids and their parents. On one hand, the ability of clonal gametogenesis occurred exclusively in hybrid germ cells, suggesting that asexual development is directly triggered by the hybrid genomic constitution of the cell. On the other hand, clonality was observed only in cells transplanted into females, suggesting that the execution of clonal development is influenced by signals from the gonadal environment and regulated by somatic factors.

Several experiments were conducted in order to develop an optimal protocol for slow-rate freezing (−1 °C/min) and short-term storage (−80 or 4 °C) of common carp ovarian tissue fragments with an emphasis on oogonial stem cells (OSCs). Dimethyl sulfoxide (Me2SO) with concentration of 1.5 M was identified as the best cryoprotectant in comparison to propylene glycol and methanol. When comparing supplementation of sugars (glucose, trehalose, sucrose) in different concentrations (0.1, 0.3, 0.5 M), glucose and trehalose in 0.3 M were identified as optimal. Short-term storage options for ovarian tissue pieces at −80 °C and 4 °C were tested as alternatives to cryopreservation and storage in liquid nitrogen. The presence of OSCs was confirmed by immunocytochemistry and viability after storage was determined by the trypan blue exclusion test. This study identified the optimal protocol for OSC cryopreservation using slow rate freezing resulting in ∼65% viability. The frozen/thawed OSCs were labelled by PKH-26 and transplanted into goldfish recipients. The success of the transplantation was confirmed by presence of fluorescent cells in the recipient gonad and later on by RT-PCR with carp dnd1 specific primers. The results of this study can facilitate long-term preservation of common carp germplasm which can be recovered in a surrogate recipient through interspecific germ cell transplantation.

Simple Summary For scientific studies on the zebrafish model, simple and routine reproductive procedures should be used to ensure stable and repeatable results. When the milt is collected, spermatozoa are spontaneously activated by urine or excrement (low osmolarity) which routinely contaminates the samples, because of the minuscule size of the fish body. Therefore, whenever milt is collected from a zebrafish for short-term milt preservation and artificial fertilization, milt must be collected into an immobilizing solution, which because of its high osmolarity stops the movement of spermatozoa and keeps the sperm immobile until fertilization. Usually, the spermatozoa showed forward movement during the 35 s period following dilution in water. The sperm concentration ranged from 0.08 to 3.52 × 109/mL with a volume from 0.1 to 2.0 μL per male. The most suitable extender proved to be E400, which allowed storage of sperm for fertilization for 6 to 12 h at a temperature of 0–2 °C. To achieve a good level of fertilization and hatchability, a test tube with a precisely defined amount of sperm with extender, eggs and activating solution proved to be the most effective. Abstract The aim of the present study was to investigate the spontaneous motility of spermatozoa and to optimize sperm collection, short-term sperm storage, and fertilization in zebrafish Danio rerio. The movement of spermatozoon in water was propagated along the flagellum at 16 s after sperm activation then damped from the end of the flagellum for 35 s and fully disappeared at 61 s after activation. For artificial fertilization, milt must be added to an immobilizing solution, which stops the movement of sperm and keeps the sperm motionless until fertilization. E400 and Kurokura as isotonic solutions were shown to be suitable extenders to store sperm for fertilization for 6 h. E400 stored sperm for 12 h at 0–2 °C. Sperm motility decreased only to 36% at 12 h post stripping for the E400 extender and to 19% for the Kurokura extender. To achieve an optimal level of fertilization and swim-up larvae rates, a test tube with a well-defined amount of 6,000,000 spermatozoa in E400 extender per 100 eggs and 100 µL of activation solution has proven to be more successful than using a Petri dish. The highest fertilization and swim-up larvae rates reached 80% and 40–60%, respectively, with milt stored for 1.5 h in the E400 extender at 0–2 °C.

DNA damage caused by exogenous or endogenous factors is a common challenge for developing fish embryos. DNA damage repair (DDR) pathways help organisms minimize adverse effects of DNA alterations. In terms of DNA repair mechanisms, sturgeons represent a particularly interesting model due to their exceptional genome plasticity. Sterlet (Acipenser ruthenus) is a relatively small species of sturgeon. The goal of this study was to assess the sensitivity of sterlet embryos to model genotoxicants (camptothecin, etoposide, and benzo[a]pyrene), and to assess DDR responses. We assessed the effects of genotoxicants on embryo survival, hatching rate, DNA fragmentation, gene expression, and phosphorylation of H2AX and ATM kinase. Exposure of sterlet embryos to 1 &micro, M benzo[a]pyrene induced low levels of DNA damage accompanied by ATM phosphorylation and xpc gene expression. Conversely, 20 &micro, M etoposide exposure induced DNA damage without activation of known DDR pathways. Effects of 10 nM camptothecin on embryo development were stage-specific, with early stages, before gastrulation, being most sensitive. Overall, this study provides foundational information for future investigation of sterlet DDR pathways.

DNA damage caused by exogenous or endogenous factors is a common challenge for developing fish embryos. DNA damage repair (DDR) pathways help organisms minimize adverse effects of DNA alterations. In terms of DNA repair mechanisms, sturgeons represent a particularly interesting model due to their exceptional genome plasticity. Sterlet (Acipenser ruthenus) is a relatively small species of sturgeon. The goal of this study was to assess the sensitivity of sterlet embryos to model genotoxicants (camptothecin, etoposide, and benzo[a]pyrene), and to assess DDR responses. We assessed the effects of genotoxicants on embryo survival, hatching rate, DNA fragmentation, gene expression, and phosphorylation of H2AX and ATM kinase. Exposure of sterlet embryos to 1 µM benzo[a]pyrene induced low levels of DNA damage accompanied by ATM phosphorylation and xpc gene expression. Conversely, 20 µM etoposide exposure induced DNA damage without activation of known DDR pathways. Effects of 10 nM camptothecin on embryo development were stage-specific, with early stages, before gastrulation, being most sensitive. Overall, this study provides foundational information for future investigation of sterlet DDR pathways.

BackgroundCommon carp is the fourth most-produced species in worldwide aquaculture. Significant efforts are invested in breeding and preservation of genetic integrity of this important species. However, maintaining carp gene bankin situcan be considered as demanding due to its big body size. Recent progress in reproductive biotechnologies in fish allows improving some unfavourable characteristics of a target species using surrogate reproduction. Germ stem cells (gamete precursors) from one species are transplanted into different surrogate species with small body size. After maturation, surrogates are producing donor-derived progeny. Efficient protocols for cryopreservation of carp male and female germ stem cells have been developed lately. Thus, the next logical goal was to assess the potential of goldfish surrogate to produce donor-derived gametes of common carp after intraperitoneal transplantation of testicular cells.ResultsHigh transplantation success was achieved when 44% of the surviving goldfish produced pure donor-derived gametes of common carp. More importantly, both viable eggs and sperm giving rise to pure common carp progeny were produced, witnessing sustainability of the presented method. Donor-derived identity of the offspring was confirmed by genotyping and typical phenotype corresponding to the donor species. Reproductive performance of chimeras was similar to goldfish controls. Assessment of gamete characteristics showed that the size of donor-derived eggs is between control carp and goldfish eggs. Interestingly, flagellum length in donor-derived spermatozoa was comparable to common carp flagellum and significantly shorter than goldfish flagellum.ConclusionsIn this study, we succeeded in the production of pure common carp progeny from surrogate goldfish recipients transplanted intraperitoneally by testicular germ cells. Here we reported production of viable eggs between most distant species up to date. Good reproductive performance of goldfish germline chimeras gives a promising prospect for further analysis about the long-term reproductive performance of surrogates, recovery of cryopreserved germ cells or production of monosex stocks. Presented technology is ready to ease needs for carp breeds preservation and their recovery using many times smaller goldfish surrogates.

Methods of induction of uniparental inheritance have been developed for model organisms as well as for species of commercial interest to fix sex-specific traits in breeding programs or to restore breeds or lines from cryopreserved sperm in aquaculture important species. Androgenesis in common carp (Cyprinus carpio) has been successfully induced by egg nucleus inactivation with γ-, X- or UV-ray irradiation but these techniques are not widely applicable for laboratory or commercial use on practical, financial and safety grounds. In the past decade a promising low-cost, low-tech cold-shock approach for successfully inducing androgenesis has been demonstrated for loach, Japanese flounder and zebrafish. The aim of the current study was to develop a cold shock methodology for application in common carp, a freshwater species of high commercial interest and a widely used model species. Gametes were collected from wild-type females (dominant green phenotype) and Koi males (recessive blonde phenotype) to enable easy identification of successful induction of androgenetic haploid progeny. A combination of different temperature treatments (0, 2, 4, 6, 8 ˚C) and different cold-shock durations (15, 30, 45, 60, 75 min.) applied shortly after gamete activation (3 s after fertilization) were initially trialed. Optimal condition for egg nucleus elimination was a cold-shock at 2 ˚C for 60 min duration where hatching rate of haploid progeny reached 35.6% and 26.26±10.19% across two attempts. Double haploid induction was then attempted with three replicates (300g egg) with following parameters 2 ˚C cold-shock for 60 min, then transferred into 20 °C and subsequently treated by a heat shock arresting first mitotic cleavage applied 40 min at 40 °C for 2 min. These combined treatments resulted in reduced fertilization and hatching rates for all replicates and low yield of progeny (1.09-1.28% in experimental incubation

Germ cells have drawn much attention as gamete precursors because of their exceptional potential for fundamental biological studies, as well as for being a potential alternative to gene resource banking for breeding and restocking purposes via surrogacy. Research has developed markers for germ cell identification, shedding light on their origin and specification mechanism. Then, pioneering studies focused on germ cell transplantation into the surrogate host with donor-derived gamete production have followed suit. The advent of transgenic technologies allowed the development of lines expressing green fluorescent protein (GFP) in their germ cells exclusively or the use of artificially synthesised mRNA conjugated with GFP to label and trace germ cells in vivo and facilitate studies on transplantation. The power of germ cell manipulation techniques was significantly enhanced by the development of a wide range of cryopreservation methods; with these approaches, valuable genetic material can be cryopreserved even during the blastula stage, as well as harvested from an adult specimen. In turn, the germ cells in the cryogenic storage can be recovered by transplantation in the surrogate host, producing donor-derived gametes after maturation. The aim of this chapter is to provide basic knowledge about a relatively novel biotechnological approach in fish reproduction and summarise performed studies on germ cell cryopreservation.

The wels catfish Silurus glanis is valuable fish for aquaculture. Its production relies mainly on artificial reproduction. One of the crucial steps determining success of the reproduction is elimination of egg stickiness after fertilization. To date, the catfish egg de-adhesion is usually carried out using proteolytic enzymes. Here, we prove a novel method based on oxidation of the egg surface by means of sodium hypochlorite. An effect of different exposure times and concentrations on the egg adhesiveness and damage was tested in the first trial. The selected concentration of sodium hypochlorite 0.3 mg · l−1 with exposure time 40 s was used for comparison with the conventical de-adhesion method using alcalase treatment. The fertilization and hatching rates reached very satisfactory outcome in both treatments (98.3 ± 0.7% vs 97.5 ± 2.2% and 86.6 ± 8.3% vs 91.3 ± 8.5% in alcalase- and sodium hypochlorite-treated embryos, respectively) without any statistical differences. Thus, the de-adhesion method using sodium hypochlorite can be recommended as a suitable method for wels catfish eggs. The method is simple, cheap, very fast, and the treated eggs are disinfected.

Nanoparticles are finding increasing applications in diagnostics, imaging and therapeutics in medicine. Iron oxide nanoparticles (IONs) have received significant interest of scientific community due to their distinctive properties. For the first time, we have delivered IONs into germ cells in any species. Our results showed that sturgeon primordial germ cells (PGCs) delivered with IONs could be detected until seven days post fertilization (dpf) under fluorescent microscope and at 22 dpf by micro-CT. Delivery of IONs into cells could be helpful for studying germ cell biology and the improvement of germ cell-based bio-technologies as isolation of PGCs using magnetic activated cell sorting or application of hyperthermia for a host sterilization purpose. Intriguingly, in our study, we did not find any toxic effects of IONs on the survival and hatching rates of sturgeon embryos when compared with embryos injected with FITC-dextran only.

We report for first time comparison of two approaches for zebrafish triploid production using cold shock and heat shock treatment. Subsequently, produced triploid zebrafish were used as a recipients for intraperitoneal transplantation of ovarian and testicular cells originating from vas:EGFP strain in order to verify their suitability for surrogate reproduction. Heat shock treatment was far more effective evaluated as success rate of triploid production and viability in comparison to cold shock. Triploids were produced with up to 100% efficiency in particular females. As expected, all triploids were males. Subsequently, germ cells transplantation revealed that triploids are suitable surrogate hosts. Production of donor-derived sperm was achieved in 23% and 16% of triploids transplanted by testicular and ovarian cells, respectively. Success of the transplantation was confirmed by positive GFP signal detected in gonads of dissected fish and stripped sperm. Germline transmission was confirmed by fertilization tests followed by PCR analysis of embryos. Reproductive success of germline chimera triploids evaluated as fertilization rate and progeny development was comparable to control groups.

Sterility and semi-sterility often occur as a result of interspecific hybridization; however, the exact cause of sterility varies across the all animal kingdom; therefore, it is in the interest of scientists to examine every hybrid species separately. In the present study, the morphology of the spermatozoa of diploid F1 hybrids of Cyprinus carpio and Carassius gibelio was examined and compared with their parents. Hybrid spermatozoa showed great variability in head and flagellum lengths. These spermatozoa were diploid and triploid often with morphological abnormalities such as multiple flagella and nuclei. They were motile within two minutes after water activation which was almost two times longer than their parental species. We also analysed the reproductive output of F1 hybrids together with parental species in both parent positions via artificial spawning. Overall, the reproductive output of the hybrid × hybrid combination was the lowest; however, the hybrids were able to produce a small number of offspring documenting the viability of the F2 generation.

Common carp (Cyprinus carpio) is one of the most cultured fish species over the world with many different breeds and plenty of published protocols for sperm cryopreservation, however, data regarding preservation of gonadal tissue and surrogate production is still missing. A protocol for freezing common carp spermatogonia was developed through varying different factors along a set of serial subsequent experiments. Among the six cryoprotectants tested, the best survival was achieved with dimethyl sulfoxide (Me2SO). In the next experiment, a wide range of cooling rates (0.5–10 °C/min) and different concentrations of Me2SO were tested resulting in the highest survival using 2 M Me2SO and cooling rate of –1 Q59 A When testing different tissue sizes and incubation times in the cryomedium, the highest viability was observed when incubating 100 mg tissue fragments for 30 min. Finally, sugar supplementation did not yield significant differences. When testing different equilibration (ES) and vitrification solutions (VS) used for needle-immersed vitrification, no significant differences were observed between the tested groups. Additionally, varied exposure time to VS did not improve the vitrification outcome where the viability was 4-fold lower than that of freezing. The functionality of cryopreserved cells was tested by interspecific transplantation into sterilized goldfish recipients. The exogenous origin of the gonads in goldfish recipients was confirmed by molecular markers and incorporation rate was over 40% in both groups at 3 months post transplantation. Results of this study can serve as an alternative way for long-term preservation of germplasm in carp which can be recovered in a surrogate recipient.

Dead end (dnd) is a germ plasm-specific maternal RNA discovered in zebrafish and then in other vertebrates. Dnd protein is essential for migration and motility of primordial germ cells (PGCs), only cells destined to transfer genetic information to offspring. PGCs arise far from somatic cells of developing gonads and they must migrate to their site of function. Migration of PGCs follows complex path by various developing tissues as their disruption impacts on the fertility. Recently, it has been found that dnd is not required for survival of PGCs and dnd-deficient zebrafish PGCs transdifferentiate into the somatic cells. In fish, targeting dnd causes removal of PGCs that ultimately affects sex differentiation. Sterility in various fish species can be achieved by knockdown or knockout of dnd. In our review, we have discussed dnd as a germ cell-specific molecular marker in fish, its interaction with miRNAs, and its use in aquaculture and fish conservation.

Common carp (Cyprinus carpio) is the fourth most-produced species in worldwide aquaculture. Significant efforts are invested in breeding and preservation of genetic integrity of this important species. However, maintaining a carp gene bank in situ can be demanding due to the large body size of the fish. Moreover, ex situ gene banking in common carp is as in other fish species limited to sperm cryopreservation. Recent progress in reproductive biotechnology in fishes has allowed the transfer of germ stem cells (gamete precursors) between individuals or even species. After maturation, the recipients are producing gametes of the donor. Surrogacy can serve as a valuable tool in germplasm banking to recover cryopreserved germ stem cells. Some unfavourable characteristics of the donor species such as long maturation time or large body size hampering its reproduction can be overcome by choosing a surrogate with more convenient characteristics. Efficient protocols for cryopreservation of common carp male and female germ stem cells have been recently developed in our laboratory. The next step has been to assess the potential of smaller goldfish (Carassius auratus) surrogates to produce donor-derived gametes of common carp after intraperitoneal transplantation of testicular cells. High transplantation success was achieved when 44% of the surviving goldfish produced pure donor-derived gametes of common carp at the age of 3 years giving rise to viable progeny. Donor-derived identity of the offspring was confirmed by genotyping and the production of a typical phenotype corresponding to the donor species. Reproductive performance of chimeras was similar to goldfish controls. Assessment of gamete characteristics showed that the size of donor-derived eggs is between control common carp and goldfish eggs. Interestingly, flagellum length in donor-derived spermatozoa was comparable to the common carp flagellum and significantly shorter than goldfish flagellum. Genotyping of the Y chromosome locus in chimeric goldfish provided a novel insight on the fate of transplanted cells, where only genotypic females were capable of producing eggs, while both genotypic males and females were capable of producing sperm. The present technology can be combined with cryopreservation procedures to ease needs for common carp breeds preservation and their recovery using many times smaller goldfish surrogates but also serve as a convenient model to study interspecific surrogacy in cyprinids.