Hybrid Embryos Research Articles

Fertile Hybrids Could Aid Coral Adaptation.

Fertile hybrids can enhance the adaptive capacity and resilience of species under stress by increasing genetic diversity within populations, masking the effects of deleterious recessive alleles, and facilitating the introgression of beneficial genetic variants into parental species. However, many hybrids are infertile. We compared the fertility of aquarium-reared F1 hybrid and purebred corals of the species Acropora loripes and Acropora kenti and examined the viability of early life stages of second-generation (F2) hybrid and back-crossed planula larvae and recruits. The F1 hybrids spawned viable gametes and the F2 hybrid and back-crossed embryos developed into planula larvae and settled to become sessile coral recruits. The F1 hybrids had greater reproductive fitness than the F1 A. loripes purebred stock in an aquarium environment based on their probability of spawning and their fertilization success in crosses using their gametes. Interspecific coral hybrids can therefore be fertile and have high reproductive fitness, which could benefit the persistence of threatened coral reefs.

Hybrid incompatibility emerges at the one-cell stage in interspecies Caenorhabditis embryos.

Intrinsic reproductive isolation occurs when genetic differences between populations disrupt the development of hybrid organisms, preventing gene flow and enforcing speciation.1-4 While prior studies have examined the genetic origins of hybrid incompatibility,5-18 the effects of incompatible factors on development remain poorly understood. Here, we investigate the mechanistic basis of hybrid incompatibility in Caenorhabditis nematodes by capitalizing on the ability of C. brenneri females to produce embryos after mating with males from several other species. Contrary to expectations, hybrid incompatibility was evident immediately after fertilization, suggesting that post-fertilization barriers to hybridization originate from physical incompatibility between sperm and oocyte-derived factors rather than from zygotic transcription, which starts after the 4-cell stage.19-22 Sperm deliver chromatin, which expands to form a pronucleus, and a pair of centrioles, which form centrosomes that attach to the sperm-derived pronucleus and signal to establish the embryo's anterior-posterior axis.23,24 In C. brenneri oocytes fertilized with C. elegans sperm, sperm pronuclear expansion was compromised, frequent centrosome detachment was observed, and cortical polarity was disrupted. Live imaging revealed that defective polar body extrusion contributes to defects in mitotic spindle morphology. C. brenneri oocytes fertilized with C. remanei or C. sp. 48 sperm showed similar defects, and their severity and frequency increased with phylogenetic distance. Defective expansion of the sperm-derived pronucleus and unreliable polar body extrusion immediately after fertilization generally underlie the inviability of hybrid embryos in this clade. These results indicate that physical mismatches between sperm and oocyte-derived structures may be a primary mechanism of hybrid incompatibility.

Successful formation of sperm cells from transplanted primordial germ cells in sterile interspecific avian recipients.

Primordial germ cells (PGCs) are stem cells, from which only gametes develop. In birds, the female sex is heterogametic, thus female gene conservation necessitates preservation of PGCs. PGC transplantation can generate germline chimeras in a host organism and develop into gametes. However, competition between host and transplanted PGCs hinder efficiency of germline chimera generation. We hypothezised that in sterile hybrid recipients with no germ cells of its own, transplanted donor PGCs may exclusively form gametes. Advantages of sterile hybrids as host for PGCs is compliant with many national regulations on genetically modified organisms and technically simpler procedure than the use of busulphan. Therefore, we investigated whether sterile interspecific hybrids may be suitable as recipients for supporting donor PGCs by injecting green fluorescent protein-labelled chicken PGCs into 3-day-old Guinea fowl and domestic fowl hybrid embryos and monitoring PGC development. The injected PGCs colonized almost 100% of the recipient gonads and produced mature spermatozoa after 44 weeks. However, gamete production in these hybrids was initiated much slower than in domestic fowls. This delay may be caused by suboptimal hormonal regulation of gametogenesis in the hybrids. Our results suggest that sterile interspecific hybrids may be suitable hosts for PGCs for efficient gamete production.

Plant Hormone Pathway Is Involved in Regulating the Embryo Development Mechanism of the Hydrangea macrophylla Hybrid.

The research is aimed to elucidate the role of plant hormones in regulating the development of hybrid embryos in Hydrangea macrophylla. Fruits from the intraspecific cross of H. macrophylla 'Otaksa' × 'Coerulea' were selected at the globular, heart, and torpedo stages of embryo development. Transcriptome sequencing and differential gene expression analysis were conducted. The results showed that fruit growth followed a single "S-shaped growth curve, with globular, heart, and torpedo embryos appearing at 30, 40, and 50 d post-pollination, respectively, and the embryo maintaining the torpedo shape from 60 to 90 d. A total of 12,933 genes was quantified across the three developmental stages, with 3359, 3803, and 3106 DEGs in the S1_vs_S2, S1_vs_S3, and S2_vs_S3 comparisons, respectively. Among these, 133 genes related to plant hormone biosynthesis and metabolism were differentially expressed, regulating the synthesis and metabolism of eight types of plant hormones, including cytokinin, auxin, gibberellin, abscisic acid, and jasmonic acid. The pathways with the most differentially expressed genes were cytokinin, auxin, and gibberellin, suggesting these hormones may play crucial roles in embryo development. In the cytokinin pathway, CKX (Hma1.2p1_0579F.1_g182670.gene, Hma1.2p1_1194F.1_g265700.gene, and NewGene_12164) genes were highly expressed during the globular embryo stage, promoting rapid cell division in the embryo. In the auxin pathway, YUC (Hma1.2p1_0271F.1_g109005.gene and Hma1.2p1_0271F.1_g109020.gene) genes were progressively up-regulated during embryo growth; the early response factor AUX/IAA (Hma1.2p1_0760F.1_g214260.gene) was down-regulated, while the later transcriptional activator ARF (NewGene_21460, NewGene_21461, and Hma1.2p1_0209F.1_g089090.gene) was up-regulated, sustaining auxin synthesis and possibly preventing the embryo from transitioning to maturity. In the gibberellin pathway, GA3ox (Hma1.2p1_0129F.1_g060100.gene) expression peaked during the heart embryo stage and then declined, while the negative regulator GA2ox (Hma1.2p1_0020F.1_g013915.gene) showed the opposite trend; and the gibberellin signaling repressor DELLA (Hma1.2p1_1054F.1_g252590.gene) increased over time, potentially inhibiting embryo development and maintaining the torpedo shape until fruit maturity. These findings preliminarily uncover the factors affecting the development of hybrid H. macrophylla embryos, laying a foundation for further research into the regulatory mechanisms of H. macrophylla hybrid embryo development.

Phylogenomics identifies parents of naturally occurring tetraploid bananas

BackgroundTriploid bananas are almost sterile. However, we succeeded in harvesting seeds from two edible triploid banana individuals (Genotype: ABB) in our conservation repository where various wild diploid bananas were also grown. The resulting rare offspring survived to seedling stages. DNA content analyses reveal that they are tetraploid. Since bananas contain maternally inherited plastids and paternally inherited mitochondria, we sequenced and assembled plastomes and mitogenomes of these seedlings to trace their hybridization history.ResultsThe coding sequences of both organellar genomic scaffolds were extracted, aligned, and concatenated for constructing phylogenetic trees. Our results suggest that these tetraploid seedlings be derived from hybridization between edible triploid bananas and wild diploid Musa balbisiana (BB) individuals. We propose that generating female triploid gametes via apomeiosis may allow the triploid maternal bananas to produce viable seeds.ConclusionsOur study suggests a practical avenue towards expanding genetic recombination and increasing genetic diversity of banana breeding programs. Further cellular studies are needed to understand the fusion and developmental processes that lead to formation of hybrid embryos in banana reproduction, polyploidization, and evolution.

Epigenetic inheritance of diet-induced and sperm-borne mitochondrial RNAs

Spermatozoa harbour a complex and environment-sensitive pool of small non-coding RNAs (sncRNAs)1, which influences offspring development and adult phenotypes1–7. Whether spermatozoa in the epididymis are directly susceptible to environmental cues is not fully understood8. Here we used two distinct paradigms of preconception acute high-fat diet to dissect epididymal versus testicular contributions to the sperm sncRNA pool and offspring health. We show that epididymal spermatozoa, but not developing germ cells, are sensitive to the environment and identify mitochondrial tRNAs (mt-tRNAs) and their fragments (mt-tsRNAs) as sperm-borne factors. In humans, mt-tsRNAs in spermatozoa correlate with body mass index, and paternal overweight at conception doubles offspring obesity risk and compromises metabolic health. Sperm sncRNA sequencing of mice mutant for genes involved in mitochondrial function, and metabolic phenotyping of their wild-type offspring, suggest that the upregulation of mt-tsRNAs is downstream of mitochondrial dysfunction. Single-embryo transcriptomics of genetically hybrid two-cell embryos demonstrated sperm-to-oocyte transfer of mt-tRNAs at fertilization and suggested their involvement in the control of early-embryo transcription. Our study supports the importance of paternal health at conception for offspring metabolism, shows that mt-tRNAs are diet-induced and sperm-borne and demonstrates, in a physiological setting, father-to-offspring transfer of sperm mitochondrial RNAs at fertilization.

Major chromosome rearrangements in intergeneric wheat × rye hybrids in compatible and incompatible crosses detected by GBS read coverage analysis

The presence of incompatibility alleles in primary amphidiploids constitutes a reproductive barrier in newly synthesized wheat-rye hybrids. To overcome this barrier, the genome stabilization process includes large-scale chromosome rearrangements. In incompatible crosses resulting in fertile amphidiploids, the elimination of one of the incompatible alleles Eml-A1 or Eml-R1b can occur already in the somatic tissue of the wheat × rye hybrid embryo. We observed that the interaction of incompatible loci Eml-A1 of wheat and Eml-R1b of rye after overcoming embryo lethality leads to hybrid sterility in primary triticale. During subsequent seed reproductions (R1, R2 or R3) most of the chromosomes of A, B, D and R subgenomes undergo rearrangement or eliminations to increase the fertility of the amphidiploid by natural selection. Genotyping-by-sequencing (GBS) coverage analysis showed that improved fertility is associated with the elimination of entire and partial chromosomes carrying factors that either cause the disruption of plant development in hybrid plants or lead to the restoration of the euploid number of chromosomes (2n = 56) in the absence of one of the incompatible alleles. Highly fertile offspring obtained in compatible and incompatible crosses can be successfully adapted for the production of triticale pre-breeding stocks.

Possibilities of using the tetraploid interspecific hybrid <i>Hordeum bulbosum</i> L. × <i>Hordeum vulgare</i> L. in obtaining new recombinant barley lines

Background. Hordeum bulbosum L. is the only wild species of the genus Hordeum, the gene pool of which is successfully used in introgressive hybridization to increase the genetic diversity of H. vulgare L. When creating introgression forms based on hybrids of H. vulgare with H. bulbosum, it is necessary to take into consideration the ploidy of the parent species, and their genetic features that determine the intensity of the process of H. bulbosum chromosomes elimination in embryogenesis. The purpose of our investigation was to study the features associated with the elimination of bulbous barley chromosomes in embryos obtained in various combinations of crosses involving the tetraploid hybrid H. bulbosum with H. vulgare for its effective use in obtaining introgression lines of H. vulgare. Material and methods. The analysis was performed on 9-15 days old embryos obtained in crosses of the tetraploid hybrid F2 (H. bulbosum A17 (4x) × H. vulgare ‘Borwina’ (4x)) (HbHbHvHv) with ‘Igri’ (2x) and ‘Borwina’ (2x) barley varieties, with H. bulbosum A17 (4x), as well as during its self-pollination. The chromosomal composition of embryos of different ages was analyzed on squashed embryo slides using genomic in situ hybridization (GISH). Results. Among the 11-15 days old embryos obtained from self-pollination of the hybrid, approximately half of the studied ones were mixoploids, while in other embryos the majority of cells contained only the genomic material of H. vulgare. The elimination process was very intensive in crosses of the hybrid with the barley varieties ‘Igri’ (2x) and ‘Borwina’ (2x), and by day 10-11 after pollination cells containing only the genomic material of H. vulgare predominated in all embryos, regardless of the direction of crossing and the variety used. Hybrid embryos with a stable chromosomal composition resulted from a cross of a tetraploid hybrid with H. bulbosum A17 (4x). Embryos with recombinant H. vulgare chromosomes carrying alien genetic material of bulbous barley were identified in all types of crosses. Conclusion. The most efficient way for the mass production of introgression lines of cultivated barley varieties based on the partially fertile hybrid F2 (H. bulbosum A17 (4x) × H. vulgare ‘Borwina’ (4x)), is the crossing with varieties of H. vulgare (2x). The progeny from such crosses will contain only cultivated barley plants, among which it is possible to identify forms with the introgression of H. bulbosum and use already the first generation from their self-pollination for selecting H. vulgare lines carrying the genetic material of bulbous barley in both homologs.

Development of regenerated plants of interspecific wheat hybrids using immature embryos as explants

Nowadays particular attention in winter wheat breeding is paid to the introduction of biotechnological methods. The culture of immature embryos in vitro is an alternative way to produce hybrids that often do not develop fully in vivo. The purpose of the current study was to estimate the ability of immature embryos of interspecific winter wheat hybrids for callus induction, morphogenesis, and plant regeneration on artificial nutrient media, as well as to select optimal conditions for obtaining full-fledged plants with all the botanical characteristics of the species. The research material included 5 hybrid combinations of interspecific crosses between durum (Triticum durum DESF.) and common (Triticum aestivum L.) winter wheat. The material was collected during one field season (2022). Immature embryos were used as explants. Culture media were according to Murashige-Skoog (MS), and they differed in the content of hormones and organic substances. There has been found that the hybrid combinations were characterized by a variety of morphogenic responses. The manifestation of callusogenesis was not high enough and averaged 22–32 % depending on the nutrient medium. Rhizogenesis averaged from 1–6.7 %, and ended with the formation of roots. The process of formation of embryoid- and hemmorizogenesis was best demonstrated by the hybrid combination No.1, and on two media. All processes of morphogenesis excepting ‘rhizogenesis’ varied greatly among hybrid combinations. The largest number of regenerated plants was obtained from hybrid combination No.1 (61 pcs., 54.5 %). After vernalization and acclimatization, 38 % of the plants survived from the number of planted embryos. The largest number of heads (79 pcs.) was formed according to hybrid combination No.1, on average 1.46 heads per plant. However, not all heads were fertile. The results of the correlation analysis made it possible to identify indicators that closely correlated with the amount of plant regeneration, namely, the high yield of embryoid- and hemmorizogenic structures ensured a high frequency of plant regeneration (r = 0.769).

Embryonic and larval development of reciprocal diploid and triploid hybrids of Caspian trout (Salmo caspius) and rainbow trout (Oncorhynchus mykiss)

The present study aims to evaluate the viability of reciprocal crosses between Caspian trout (Salmo caspius) and rainbow trout (Oncorhynchus mykiss), and the effect of triploidization on embryonic development, growth and survival of hybrid embryos and larvae. This experiment included six groups of crosses: (1) pure O. mykiss (2nR); (2) pure S. caspius (2nC); (3) diploid and (4) triploid ♀O. mykiss×♂S. caspius hybrids (2nR × C and 3nR × C); (5) diploid and (6) triploid ♀S. caspius×♂O. mykiss hybrids (2nC × R and 3nC × R). Triploidy was induced by a thermal shock after fertilization. Fertilization, hatching, and embryonic development were studied. Hatching rate was more than 90% in 2nR and 2nC, 35% in 2nR × C group, 45% in 3nR × C group, whereas all embryos died before the eyed stage in 2nC × R and 3nC × R groups. Chromosome number ranged between 57 and 61 and 78–81 in 2nR and 2nC groups, 69–71 in 2nR × C and 99–103 in 3nR × C. The time before blastopore closure was the most important point in the embryonic phases when most deaths occurred in all groups. Hatching occurred earlier in 2nR and 3nR × C groups than in 2nC group. The highest final weight and specific growth rate were observed in 2nR and 3nR × C groups. We have demonstrated that triploidy induction can increase the survival rate of distant hybrid (♀O. mykiss×♂S. caspius) and identified the key embryonic stages that could help introduce this hybrid into aquaculture. The hybrid could be easily commercialized if they show significant improvement in growth, survival and disease resistance.

Antagonistic RALF peptides control an intergeneric hybridization barrier on Brassicaceae stigmas

Pollen-pistil interactions establish interspecific/intergeneric pre-zygotic hybridization barriers in plants. The rejection of undesired pollen at the stigma is crucial to avoid outcrossing but can be overcome with the support of mentor pollen. The mechanisms underlying this hybridization barrier are largely unknown. Here, in Arabidopsis, we demonstrate that receptor-like kinases FERONIA/CURVY1/ANJEA/HERCULES RECEPTOR KINASE 1 and cell wall proteins LRX3/4/5 interact on papilla cell surfaces with autocrine stigmatic RALF1/22/23/33 peptide ligands (sRALFs) to establish a lock that blocks the penetration of undesired pollen tubes. Compatible pollen-derived RALF10/11/12/13/25/26/30 peptides (pRALFs) act as a key, outcompeting sRALFs and enabling pollen tube penetration. By treating Arabidopsis stigmas with synthetic pRALFs, we unlock the barrier, facilitating pollen tube penetration from distantly related Brassicaceae species and resulting in interspecific/intergeneric hybrid embryo formation. Therefore, we uncover a "lock-and-key" system governing the hybridization breadth of interspecific/intergeneric crosses in Brassicaceae. Manipulating this system holds promise for facilitating broad hybridization in crops.

Evolutionary conservation of embryonic DNA methylome remodelling in distantly related teleost species.

Methylation of cytosines in the CG context (mCG) is the most abundant DNA modification in vertebrates that plays crucial roles in cellular differentiation and identity. After fertilization, DNA methylation patterns inherited from parental gametes are remodelled into a state compatible with embryogenesis. In mammals, this is achieved through the global erasure and re-establishment of DNA methylation patterns. However, in non-mammalian vertebrates like zebrafish, no global erasure has been observed. To investigate the evolutionary conservation and divergence of DNA methylation remodelling in teleosts, we generated base resolution DNA methylome datasets of developing medaka and medaka-zebrafish hybrid embryos. In contrast to previous reports, we show that medaka display comparable DNA methylome dynamics to zebrafish with high gametic mCG levels (sperm: ∼90%; egg: ∼75%), and adoption of a paternal-like methylome during early embryogenesis, with no signs of prior DNA methylation erasure. We also demonstrate that non-canonical DNA methylation (mCH) reprogramming at TGCT tandem repeats is a conserved feature of teleost embryogenesis. Lastly, we find remarkable evolutionary conservation of DNA methylation remodelling patterns in medaka-zebrafish hybrids, indicative of compatible DNA methylation maintenance machinery in far-related teleost species. Overall, these results suggest strong evolutionary conservation of DNA methylation remodelling pathways in teleosts, which is distinct from the global DNA methylome erasure and reestablishment observed in mammals.

Standardization of a Sex-Sorting Protocol for Stallion Spermatozoa by Means of Absolute RT-qPCR.

Sperm sexing is a technology that can generate great economic benefits in the animal production sector. Techniques such as sex-sorting promise over 90% accuracy in sperm sexing. However, for the correct standardization of the technique, some laboratory methodologies are required. The present manuscript describes in detail a standardized equine sperm sex-sorting protocol using an absolute qPCR-based methodology. Furthermore, the results of absolute qPCR were implemented and validated by generating equine/bovine heterologous embryos by intracytoplasmic sperm injection (ICSI) of presumably sexed equine spermatozoa into bovine oocytes using a piezoelectric system (Piezo-ICSI). Our results indicated that equine sex-sorting spermatozoa had a 97% and 94% certainty for X and Y sperm, respectively, while presumptive female and male equine/bovine hybrid embryos, generated by Piezo-ICSI, had an accuracy of 92% with respect to the desired sex. Therefore, it is concluded that the presented methodology is a reliable, cost-effective, and relatively simple option for standardizing sex-sorting of equine spermatozoa. This is supported by the results of the correct sexing of Piezo-ICSI heterologous embryos generated with the sexed spermatozoa, validating the correct sexing and viability of these gametes.

Understanding and overcoming hybrid lethality in seed and seedling stages as barriers to hybridization and gene flow.

Hybrid lethality is a type of reproductive isolation barrier observed in two developmental stages, hybrid embryos (hybrid seeds) and hybrid seedlings. Hybrid lethality has been reported in many plant species and limits distant hybridization breeding including interspecific and intergeneric hybridization, which increases genetic diversity and contributes to produce new germplasm for agricultural purposes. Recent studies have provided molecular and genetic evidence suggesting that underlying causes of hybrid lethality involve epistatic interaction of one or more loci, as hypothesized by the Bateson-Dobzhansky-Muller model, and effective ploidy or endosperm balance number. In this review, we focus on the similarities and differences between hybrid seed lethality and hybrid seedling lethality, as well as methods of recovering seed/seedling activity to circumvent hybrid lethality. Current knowledge summarized in our article will provides new insights into the mechanisms of hybrid lethality and effective methods for circumventing hybrid lethality.

Transcriptome Analysis Reveals the Role of Phytohormones in the Distant Hybridization of Peony Embryo Abortion

Peonies are significant ornamental plants that are primarily propagated through distant cross-breeding to create new varieties. However, hybrid failure is a critical issue that impedes the advancement of breeding. Numerous studies have demonstrated that endogenous hormones in the seed embryo constitute a significant factor in embryo failure. Nevertheless, it is still unknown how plant hormones control the development of peony embryos at the molecular level. In this study, we characterized the endogenous hormone levels in peony seeds of hybrid-aborted, hybrid-normal, and self-crossing normal after 26 days of pollination (DAP). Our findings show that the hybrid-aborted embryos had significantly higher amounts of ABA, IAA, and GA. In addition, the ratio of GA+IAA/ABA was lower than that of the hybrid-normal seeds and higher than that of the self-crossing normal seeds. To further investigate the mechanism of hormone control on peony embryo development, we conducted a transcriptome sequencing analysis of the three seed types. Results revealed that differentially expressed genes involved in phytohormone metabolism and signal transduction significantly enriched the aborted embryos. Furthermore, we examined the expression levels of six hormones in different seeds and used the Gene Common Expression Trend analysis to analyze genes highly correlated with phytohormone in the KEGG pathway. We used protein interaction networks to explore the interactions between proteins in the hormone pathway in aborted embryos. Then we identified key genes and transcription factors (TFs) such as Abscisic acid-insensitive 5 (ABI5), Auxin Response Factor 5 (ARF5), Gibberellin Insensitive Dwarf 1 (GID1), Arabidopsis Response Regulator4 (ARR4), Jasmonate-zim-domain protein 1 (JAZ1), Brassinazole-resistant 1 (BZR1), etc., whose functions require further investigation. Our findings establish a foundation for the metabolic regulation of peony hybrid embryo abortion via networks regulating phytohormone signaling. However, further research is needed to determine the exact mechanisms by which hormones regulate peony embryo development and to explore new methods for improving the success rate of hybridization.

Success of embryo transfer of hybrid embryos in Old World camelids: A first study.

This is the first study to test the embryo transfer (ET) of hybrid embryos in Old World camelids and produces a live calf from a dromedary recipient. Hybrid embryos were collected from 7 dromedary and 10 Bactrian donors, with or without ovarian super-stimulation, and transferred to dromedary recipients. Pregnancy diagnosis was conducted on Day 10 post-ET by using the progesterone-ELISA test and trans-rectal ultrasonography at 1 and 2 months of gestation. The date of abortion, stillbirth, or normal calving for each pregnant recipient was recorded. Without ovarian super-stimulation, two and one recipients were pregnant at 10 days post-ET, from ♂ Bactrian X ♀ dromedary and ♂ dromedary X ♀ Bactrian, respectively. While at 2 months of gestation, only one recipient was diagnosed pregnant from ♂ Bactrian X ♀ dromedary. Response to ovarian super-stimulation was successful in all 4 of the tested dromedary donors and in 8 out of 10 Bactrian donors. Additionally, 4 super-stimulated Bactrian donors (40%) showed failure of ovulation. The number of super-stimulated developed follicles and recovered embryos was higher in dromedary donors compared to Bactrian donors. Ten and two recipients were diagnosed pregnant at 10 days post-ET for ♂ Bactrian X ♀ dromedary and ♂ dromedary X ♀ Bactrian, respectively. At 2 months of gestation, the number of pregnant recipients from ♂ Bactrian X ♀ dromedary was reduced to eight, while the two pregnant recipients from ♂ dromedary X ♀ Bactrian remained pregnant. Total early pregnancy loss at 2 months gestation for all transferred hybrid embryos, obtained with or without ovarian super-stimulation, was 4/15 (26.6%). One healthy male calf was born from a recipient, with a total gestation period of 383 days, that received an embryo from a Bactrian male and a dromedary donor. Stillbirth was observed in six cases after 10.5-12 months of gestation and three cases aborted between 7 and 9 months of gestation due to trypanosomiasis. In conclusion, ET of hybrid embryos in Old World camelids is successful. However, further studies are required to improve the outcome of this technology to be used for meat and milk production in camels.

Acquisition of fertilization competence in guinea pig spermatozoa under different capacitation protocols

Guinea pig in vitro fertilization (IVF) are poorly developed due to the limited accessibility to oocytes and the lack of an efficient method of sperm capacitation. Thus, we aimed to evaluate different capacitation protocols that we validated through sperm analysis and using heterologous (He) IVF with zona-intact bovine oocytes. Spermatozoa of guinea pigs were collected and processed separately by 4 different protocols: A) Spermatozoa were obtained by flushing the lumen of one cauda epididymis and incubated in a minimal culture medium (MCM); B) One epididymis was placed in a prewarmed of M2 medium and gently minced with fine scissors. Spermatozoa were incubated in a modified human tubal fluid medium (HTF). In both protocols, the spermatozoa were capacitated at 37 °C under an atmosphere of 5% CO2 for 2 h. In the protocols C and D, the spermatozoa were collected by flushing the lumen of the cauda epididymis and selected by commercial density gradient Bovipure® (Nidacon Laboratories AB, Göthenborg, Sweden), according to the manufacturer's instructions. Then for Protocol C) spermatozoa were incubated in MCM medium supplemented with 10 mg/mL heparin (MCM-Hep); while for Protocol D) spermatozoa were incubated in FERT medium supplemented 10 mg/mL heparin (FERT-Hep). Incubation of C and D protocols were performed at 38.5 °C under an atmosphere of 5% CO2 for 2 h. Capacitation protocols C and D showed a higher percentage of viability, total and hyperactive-like motility, and acrosome reaction compared to protocols A and B. For this reason, protocols C and D were used for further He-IVF analysis. Guinea pig sperm and matured zona-intact bovine oocytes were co-incubated at 5% CO2 and 38.5 °C. Sperm-oocyte interaction was assessed at 2.5 h post-insemination (hpi) and pronuclear formation (PrF) were evaluated at 18, 20, 22, 24 and 26 hpi, while the cleavage rate was evaluated at 48 hpi. In protocol D, PrF was significantly higher than in protocol C (P ≤ 0.05) at every time point evaluated. Also, the cleavage rate at 48 hpi was higher (P ≤ 0.05) in He-IVF protocol D (69.8 ± 1.7%) compared to He-IVF protocol C (49.1 ± 1.1%). In conclusion, we determined the most adequate sperm capacitation conditions for guinea pig that allow zona-intact bovine oocyte penetration and lead to hybrid embryo formation, suggesting that these conditions could be optimal to develop IVF in guinea pigs.

Embryonic Development and Survival of Siberian Sturgeon × Russian Sturgeon (Acipenser baerii × Acipenser gueldenstaedtii) Hybrids Cultured in a RAS System.

The aim of this study was to describe the selected stages of embryonic development in Siberian sturgeon (Acipenser baerii) and hybrids of Siberian sturgeons and Russian sturgeons (Acipenser baerii × Acipenser gueldenstaedtii). For this purpose, embryos representing nine distinct developmental stages (stage 1-2.0 hpf, stage 2-5.5 hpf, stage 3-13.0 hpf, stage 4-20.0 hpf, stage 5-24.0 hpf, stage 6-26.0 hpf, stage 7-35.0 hpf, stage 8-55.0 hpf, and, stage 9-160.0 hpf; hpf-hours postfertilization) were sampled from each group (group A, group B, and group C) during incubation. Stages of embryogenesis were identified based on a 30-point scale of embryonic development in sturgeons. A total of 13 developmental stages were identified, including early cleavage, blastula formation, early and late gastrulation, onset of neurulation, beginning of organogenesis, and prelarvae. During gastrulation, the survival of hybrid embryos was highest in group B (93.8%) and lowest in group A (86.7%). Embryonic deformation was not observed during experimental incubation. The archived data relating to the embryonic development of Siberian sturgeon × Russian sturgeon hybrids could be applied to identify the individual stages of embryogenesis in hybrid sturgeons during egg incubation.

Understanding chromosome elimination in hybrid embryo cells for innovations in plant breeding

Understanding chromosome elimination in hybrid embryo cells for innovations in plant breeding

The use of maize haploidy inducers as a tool in agricultural plant biotechnology.

The discovery of the ability of some mutations to stimulate haploidy during hybridization made it possible to create one of the most promising and sought-after trends in the f ield of reproductive biology. Haploid inducers created on their basis are capable of increasing the frequency of haploidy up to 15 %. The improvement of the existing haploid inducer lines and the search for new genes that contribute to a high frequency of haploidy are underway. Along with these studies, the f ield of application of haploid inducers in genetics and plant breeding is expanding. Haploid inducers carrying R1-nj genes for anthocyanin pigmentation of the seed and embryo are able not only to mark the hybrid embryo and identify haploid genotypes, but also to detect genes that suppress the anthocyanin color of the grain, like C1-I, C2-Idf, and In1-D. Depending on their quantity, the phenotypic manifestation of the gene in the seed varies. Haploidy is widely used for accelerating hybrid breeding and obtaining both new maize lines with improved traits and their sterile counterparts. By introducing certain genes into the genome of the improved line, breeders can use the doubled haploid (DH) breeding technology to accelerate the creation of pure lines carrying the desired gene. Haploid inducer maize lines and their tetraploid analogs are used in the selection of rediploid maize lines by their resynthesis from tetraploid genotypes. In 2019, Syngenta Company synthesized a haploid inducer maize line carrying a CRISPR/cas construct capable of simultaneously stimulating haploidy and editing the genome at a specif ied DNA site. Thanks to this technology, it became possible to improve haploid inducers by introducing various CRISPR/cas constructs into the haploid inducer genome for editing any DNA site. Maize haploid inducers are widely used in doubled haploid wheat breeding. The f irst experiments showed that the most effective haploid inducer for stimulating haploidy in wheat is maize pollen. Researchers are intensively searching for other ways of using maize haploid inducers in plant breeding.