Red Crucian Carp Research Articles

Localization of RNA Pol II CTD (S5) and Transcriptome Analysis of Testis in Diploid and Tetraploid Hybrids of Red Crucian Carp (♀) × Common Carp (♂).

Polyploidy occurs naturally in fish; however, the appearance of these species is an occasional and gradual process, which makes it difficult to trace the changes in phenotypes, genotypes, and regulation of gene expression. The allotetraploid hybrids (4nAT) of red crucian carp (RCC; ♀) × common carp (CC; ♂) generated from interspecies crossing are a good model to investigate the initial changes after allopolyploidization. In the present study, we focused on the changes in the active sites of the testicular transcriptome of the allotetraploid by localization of RNA Pol II CTD YSPTSPS (phospho S5) using immunofluorescence and RNA-seq data via bioinformatic analysis. The results showed that there was no significant difference in signal counts of the RNA Pol II CTD (S5) between the different types of fish at the same stages, including RCC, CC, 2nF1, and 4nAT, which means that the number of transcriptionally active sites on germ cell chromosomes was not affected by the increase in chromosome number. Similarly, RNA-seq analysis indicated that in the levels of chromosomes and 10-kb regions in the genome, there were no significant changes in the highly active sites in RCC, 2nF1, and 4nAT. These findings suggest that at the beginning of tetraploid origin, the active transcriptome site of 4nAT in the testis was conserved in the regions of the genome compared to that in RCC and 2nF1. In conclusion, 4nAT shared a similar gene expression model in the regions of the genome with RCC and 2nF1 with significantly different expression levels.

Elevated expression of inhibin α gene in sterile allotriploid crucian carp

Inhibin and Activin, belong to the transforming growth factor β superfamily (TGF-β), which associate with the regulation of the reproductive process by the modulation of the hypothalamic-pituitary-gonad (HPG) axis. In this study, we reported the molecular cloning and tissue expression of inhibin α in allotriploid crucian carp and its parent- diploid red crucian carp. The full-length cDNA of inhibin α were respectively 1632 bp and 1642 bp in allotriploids and diploids, which both consisted of a 1044 bp open reading frame (ORF) encoding 347 amino acids. Real-time quantitative PCR (RT-qPCR) showed that allotriploids and diploids had significant expression of inhibin α in testis and ovary, and the expression of inhibin α in the gonads of allotriploids was higher than that of diploids. The immunohistochemistry indicated that the ovarian development of allotriploids was abnormal, and the expression of Inhibin α in the ovary of allotriploids was higher than that of diploids. Results of co-immunoprecitation (co-IP) demonstrated that the Inhibin α and Activin βA, Inhibin α and Activin βB can form dimers. These findings suggested that the elevated expression of inhibin α and the competitive binding of Inhibin α subunit with Activin β subunits in allotriploids may be releted to the sterility of allotriploids. Furthermore, these results will facilitate the investigation of reproduction characteristics in allotriploids and provide theoretical basis for the study of polyploid breeding in the future.

A novel NK-lysin in hybrid crucian carp can exhibit cytotoxic activity in fish cells and confer protection against Aeromonas hydrophila infection in comparison with Carassius cuvieri and Carassius auratus red var

NK-lysin, an effector of natural killer (NK) cells and cytotoxic T lymphocytes (CTLs), not only exhibits cytotoxic effect in fish cells, but also participates in the immune defense against pathogenic infection. In this study, ORF sequences of RCC-NK-lysin, WCC-NK-lysin and WR-NK-lysin were 369 bp. Tissue-specific analysis revealed that the highest expressions of RCC-NK-lysin and WCC-NK-lysin were observed in gill, while the peaked level of WR-NK-lysin mRNA was observed in spleen. A. hydrophila infection sharply increased RCC-NK-lysin, WCC-NK-lysin and WR-NK-lysin mRNA expression in liver, trunk kidney and spleen. In addition, elevated levels of NK-lysin mRNA were observed in cultured fin cell lines of red crucian carp (RCC), white crucian carp (WCC) and their hybrid offspring (WR) after Lipopolysaccharide (LPS) challenge. RCC-NK-lysin, WCC-NK-lysin and WR-NK-lysin exerted regulatory roles in inducing ROS generation, modulating mitochondrial membrane potential, decreasing fish cell viability and antagonizing survival signalings, respectively. RCC/WCC/WR-NK-lysin-overexpressing fish could up-regulate expressions of inflammatory cytokines and decrease bacterial loads in spleen. These results indicated that NK-lysin in hybrid fish contained close sequence similarity to those of its parents, possessing the capacities of cytotoxicity and immune defense against bacterial infection.

A new type of triploid fish derived from the diploid hybrid crucian carp (♀) × autotetraploid fish (♂)

Based on the allodiploid lineage (WR, 2n ​= ​100, F1–F5) of white crucian carp (Carassius cuvieri, WCC, 2n ​= ​100) (♀) ​× ​red crucian carp (Carassius auratus red var., RCC, 2n ​= ​100) (♂) and the diploid hybrid fish (WR-II, 2n ​= ​100) derived from WR-F1 (♀) ​× ​WCC (♂), in this research, we obtain a new type of triploid fish (WR-III) by the distant hybridization of WR-II (♀) and autotetraploid fish (4nAU, 4n ​= ​200) (♂). We investigate the morphological traits, chromosomal numbers, DNA content, growth rates and 5S rDNA in the WR-III. The results show that the WR-III is triploid fish with 150 chromosomes and they have no barbel and have a gray body color. All the measurable traits of WR-III show hybrid characteristics; and there is a significant difference in the body length/body height between WR-III and 4nAU, which indicating that WR-III has a smaller head and higher body compared with 4nAU; furthermore, the WR-III has a fast growth rate, the average body weights of eight months old WR-III is 487g. The 5s rDNA of the WR-III inherits the 5s rDNA of its parents and has some mutant bases or chimeric sequences, which revealed heredity, variability and heterozygosity in WR-III. The successful production of this triploid fish is of great significance in fish genetic breeding.

Integration of miRNA-mRNA co-expression network reveals potential regulation of miRNAs in hypothalamus from sterile triploid crucian carp

The sterile triploid crucian carp (TC) has growth superiority which is generated from crossing between diploid red crucian carp (Carassius carassius red var. ♀) (RC) and tetraploid hybrid linage [derived from RC (♀) × common carp (Cyprinus carpio, ♂) (CC)]. Hypothalamus is an important control center for hormone releasing participating in gonadal development. In the present study, the microRNA (miRNA) alternations in the hypothalamus from RC, CC and TC were analyzed. A total of 143 known miRNAs and 519 novel miRNAs were identified from the hypothalamus. Thereinto, miR-7a, miR-128 and miR-9-3p were the most abundant miRNAs accounting for 13.37%, 12.01% and 11.15% of all the miRNA reads, respectively. A total of 109, 208 and 228 differentially expressed miRNAs (DEMs) were identified from RC vs TC, CC vs TC and CC vs RC, respectively. For these DEMs, 46 miRNA-mRNA pairs were identified by miRNA target prediction. In addition, using weighted gene co-expression network analysis (WGCNA), 17 modules of co-expressed miRNAs and mRNA were constructed and yellow module had the highest correlation with the sterile triploid. A total of 107 novel miRNAs, 29 conserved miRNAs and 1343 genes were contained in yellow module and metabolic processes, biosynthetic processes, chromosome organization, cytoskeleton organization, gene expression and axis specification were enriched GO terms in this module. The top three hub miRNAs were let-7a, miR-100 and miR-122 which were correlated with 1086, 1172 and 1222 genes, respectively. These results indicated that the hub miRNAs and their regulated gene network play a central role in controlling endocrine secretion and gamete maturation. In addition, the abnormal expression of these miRNAs contributes to the sterility of TC. The present evidence provides insights into the miRNA and gene constructed network of sterility in triploid fish.

Effect of Lipopolysaccharide (LPS) stimulation on apoptotic process and oxidative stress in fibroblast cell of hybrid crucian carp compared with those of Carassius cuvieri and Carassius auratus red var

Bacterial LPS is a heat-stable endotoxin and wall components of gram negative bacteria, which can exhibit a toxicological effect on physiology and biochemical activities of fish. In this study, we investigated the effect of LPS exposure on cell viability, oxidative stress, caspase activity and immune-related gene expressions in cultured fin cell lines of red crucian carp, white crucian carp and their hybrid offspring. LPS stimulation could reduce fish cell viability, whereas gene expression levels and promoter activities in inflammatory signals increased dramatically. Moreover, enhanced levels of intracellular oxidative stress and decreased levels of mitochondrial membrane potential (MMP) were observed in LPS-induced fish cells. N-Acetyl-L-cysteine (NAC) could alleviate LPS-stimulated reactive oxygen species (ROS) generation and caspase-3 activity in fish cells. These results suggested that ROS-mediated cytotoxic stress was involved in LPS-induced inflammation and mitochondrial damage in cultured fish cells.

Variations in the Mitochondrial Genome of a Goldfish-Like Hybrid [Koi Carp (♀) × Blunt Snout Bream (♂)] Indicate Paternal Leakage.

Previously, a homodiploid goldfish-like fish (2n = 100; GF-L) was spontaneously generated by self-crossing a homodiploid red crucian carp-like fish (2n = 100; RCC-L), which was in turn produced via the distant hybridization of female koi carp (Cyprinus carpio haematopterus, KOC, 2n = 100) and male blunt snout bream (Megalobrama amblycephala, BSB, 2n = 48). The phenotypes and genotypes of RCC-L and GF-L differed from those of the parental species but were similar to diploid red crucian carp (2n = 100; RCC) and goldfish (2n = 100; GF), respectively. We sequenced the complete mitochondrial DNAs (mtDNAs) of the KOC, BSB, RCC-L, GF-L, and subsequent generations produced by self-crossing [the self-mating offspring of RCC-L (RCC-L-F2) to the self-mating offspring of RCC-L-F2 (RCC-L-F3) and the self-mating offspring of GF-L (GF-L-F2)]. Paternal mtDNA fragments were stably embedded in the mtDNAs of both lineages, forming chimeric DNA fragments. In addition to these chimeras, several nucleotide positions in the RCC-L and GF-L lineages differed from the parental bases, and were instead identical with RCC and GF, respectively. Moreover, RCC-L and GF-L mtDNA organization and nucleotide composition were more similar to those of RCC and GF, respectively, compared to parental mtDNA. Finally, phylogenetic analyses indicated that RCC-L and GF-L clustered with RCC and GF, not with the parental species. The molecular dating time shows that the divergence time of KOC and GF was about 21.26 Mya [95% highest posterior density (HPD): 24.41–16.67 Mya], which fell within the period of recent. The heritable chimeric DNA fragments and mutant loci identified in the mtDNA of the RCC-L and GF-L lineages provided important evidence that hybridizations might lead to changes in the mtDNA and the subsequent generation of new lineages. Our findings also demonstrated for the first time that the paternal mtDNA was transmitted into the mtDNA of homodiploid lineages (RCC-L and GF-L), which provided evidence that paternal DNA plays a role in inherited mtDNA. These evolutionary analyses in mtDNA suggest that GF might have diverged from RCC after RCC diverged from koi carp.

Comparative analyses of hypothalamus transcriptomes reveal fertility-, growth-, and immune-related genes and signal pathways in different ploidy cyprinid fish

Triploid crucian carp (TCC) is obtained by hybridization of female diploid red crucian carp (Carassius auratus red var., RCC) and male allotetraploid hybrids. In this study, high-throughput sequencing was used to conduct the transcriptome analysis of the female hypothalamus of diploid RCC, diploid common carp (Cyprinus carpio L., CC) and TCC. The key functional expression genes of the hypothalamus were obtained through functional gene annotation and differential gene expression screening. A total of 71.56 G data and 47,572 genes were obtained through sequencing and genome mapping, respectively. The Fuzzy Analysis Clustering assigned the differentially expressed genes (DEGs) into eight groups, two of which, overdominance expression (6005, 12.62%) and underdominance expression (3849, 8.09%) in TCC were further studied. KEGG enrichment analysis showed that the DEGs in overdominance were mainly enriched in four pathways. The expression of several fertility-related genes was lower levels in TCC, whereas the expression of several growth-related genes and immune-related genes was higher levels in TCC. Besides, 15 DEGs were verified by quantitative real-time PCR (qPCR). The present study can provide a reference for breeding sterility, fast-growth, and disease-resistant varieties by distant hybridization.

Chimeric ferritin H in hybrid crucian carp exhibits a similar down-regulation in lipopolysaccharide-induced NF-κB inflammatory signal in comparison with Carassius cuvieri and Carassius auratus red var

Ferritin H can participate in the regulation of teleostean immunity. ORF sequences of RCC/WCC/WR-ferritin H were 609 bp, while WR-ferritin H gene possessed chimeric fragments or offspring-specific mutations. In order to elucidate regulation of immune-related signal transduction, three fibroblast-like cell lines derived from caudal fin of red crucian carp (RCC), white crucian carp (WCC) and their hybrid offspring (WR) were characterized and designated as RCCFCs, WCCFCs and WRFCs. A sharp increase of ferritin H mRNA was observed in RCCFCs, WCCFCs and WRFCs following lipopolysaccharide (LPS) challenge. Overexpression of RCC/WCC/WR-ferritin H can decrease MyD88-IRAK4 signal and antagonize NF-κB, TNFα promoter activity in RCCFCs, WCCFCs and WRFCs, respectively. These results indicated that ferritin H in hybrid offspring harbors highly-conserved domains with a close sequence similarity to those of its parents, playing a regulatory role in inflammatory signals.

Genetic Variation in an Experimental Goldfish Derived From Hybridization.

Owning to the extreme difficulty in identifying the primary generation (G0), the common ancestor of various twin-tail goldfish strains remains unclear. However, several authors have hypothesized that this ancestor may have been the crucian carp (Carassius auratus). Previously, we generated an experimental hybrid goldfish (EG) from the interspecific hybridization of red crucian carp (Carassius auratus ♀, RCC) × common carp (Cyprinus carpio ♂, CC). Unlike either parent, EG possessed twin caudal fins similar to those of natural goldfish (Carassius auratus, NG). The genetic characteristics of EG, as well as the mechanisms underlying its formation, are largely unknown. Here, we identified the genetic variation in the chordin gene that was associated with the formation of the twin-tail phenotype in EG: a stop codon mutation at the 127th amino acid. Furthermore, simple sequence repeat (SSR) genotyping indicated that, among the six alleles, all of the EG alleles were also present in female parent (RCC), but alleles specific to the male parent (CC) were completely lost. At some loci, EG and NG alleles differed, showing that these morphologically similar goldfish were genetically dissimilar. Collectively, our results demonstrated that genetic variations and differentiation contributed to the changes of morphological characteristics in hybrid offspring. This analysis of genetic variation in EG sheds new light on the common ancestor of NG, as well as on the role of hybridization and artificial breeding in NG speciation.

Species and tissue specific analysis based on quantitative proteomics from allotetraploid and the parents

Allotetraploids play central roles in the field of polyploid breeding of freshwater fishes. The molecular basis underlying distant hybridization and individual differences between allotetraploids and their parents is largely unknown. In this study, we performed quantitative proteomics profiling in gonad and liver tissues between allotetraploid and its parents Red Crucian Carp (♀) and Common Carp (♂) respectively. Thousands of proteins were identified and quantified. Species and tissue specific analysis revealed that a large number of causal proteins are specifically regulated in gonad or liver tissue in allotetraploid and its parents respectively. Subsequently, integrative bioinformatics analyses including functional enrichment, pathway and network analyses were conducted. The results suggested a series of gonad and liver specifically regulated proteins such as LSM3, LSM7, PABPC1B and ALDH3A1, EHHADHB, ACAT2 play crucial roles in reproduction-related and metabolism-related pathways including “DNA replication”, “Spliceosome” and “Metabolic pathways”, “Biosynthesis of antibiotics”. Meanwhile, species specifically regulated proteins such as FMR1, MAO are involved in “RNA transport”, “Glycine, serine and threonine metabolism”. Herein, we established the first comprehensive proteomics knowledgebase for particular freshwater fishes. It may shed light on the molecular mechanisms underlying polyploidy breeding and individual differences and serve as an indispensable reference for further studies. SignificanceThe molecular basis underlying distant hybridization and individual differences between allotetraploid and their parents Red Crucian Carp and Common Carp is largely unknown. Quantitative proteomics profiling integrated with multiple bioinformatics analysis revealed that a large number of causal proteins are specifically expressed in gonad or liver tissue in allotetraploid and its parents. Herein, we established the first comprehensive proteomics knowledgebase for particular freshwater fishes. It may shed light on the molecular mechanisms underlying polyploidy breeding and individual differences and serve as an indispensable reference for further associated studies.

Upregulation of TDRD1 Promotes the Sexual Maturation in Allotetraploids Hybridized from Red Crucian Carp (Carassius auratus Red var) (♀) and Common Carp (Cyprinus carpio L) (♂).

Allotetraploid hybrids of Red Crucian Carp (Carassius auratus red var, abbreviated as RCC) (♀) and common Carp (Cyprinus carpio L, abbreviated as CC) (♂) is a species produced by distant hybridization. In this study, SWATH mass spectrometry (SWATH-MS) was applied for quantitative proteomics profiling of gonad tissues of allotetraploids and their parents RCC (♀) and CC (♂). A total of 2338 unique proteins were identified in our proteomic profiling by SWATH-MS. Gene enrichment and network analysis based on differentially expressed proteins revealed some metabolic enzymes involved in gonad growth and development of allotetraploids. Especially, the upregulated gene TDRD1 (ratio = 2.59, p = 0.02) in allotetraploids plays a significant role in spermatogenesis, which could speed up the sexual maturation during the process of gonad growth and development. Protein-protein interaction and pathway analysis further suggested that TDRD1 served as a hub protein in metabolic-related pathways and networks. It could regulate gonad development via regulating metabolic pathways in a synergistic way with surrounding regulatory factors including CS, GPIA, PGK1, and IDH2. In addition, TDRD1 could directly regulate spermatogenesis in conjunction with PIWIL1, PIWIL2, and VASA. Quantitative proteomics integrated with network analysis explored the molecular mechanism in that TDRD1 regulated sexual maturation, development, and growth of allotetraploids in a synergetic way with metabolic genes and pathways.

3nLcn2, a teleost lipocalin 2 that possesses antimicrobial activity and inhibits bacterial infection in triploid crucian carp

Lipocalin 2 (Lcn2) has been identified in mammals, however, the in vivo function of fish Lcn2 is essentially unknown. Triploid crucian carp (3n = 150) of red crucian carp (female, 2n = 100) and allotetraploid (male, 4n = 200) shows better resistance to pathogenic infections. To elucidate the antimicrobial mechanism of triploid crucian carp, we examined the function of a novel Lcn2 from triploid crucian carp (3nLcn2). 3nLcn2 is 183 residues in length and contains a conserved lipocalin domain. Quantitative real time reverse transcription PCR (qRT-PCR) analysis showed that 3nLcn2 expression occurred in multiple tissues and was upregulated by bacterial infection in a time-dependent manner. We found that purified recombinant 3nLcn2 (r3nLcn2) exerted bactericidal activity to Aeromonas hydrophila and Escherichia coli. qRT-PCR detected increased expression of pro-inflammatory cytokines and tight junctions in fish with 3nLcn2 overexpression. Fish administered with 3nLcn2 exhibited enhanced intestinal barrier and resistance against bacterial infection. These results provide the first evidence that 3nLcn2 is a functional lipocalin with antimicrobial activity and plays a positive role in the immune defense during bacterial infection.

Rapid Genomic and Epigenetic Alterations in Gynogenetic Carassius auratus Red Var. Derived from Distant Hybridization.

Gynogenesis is an important reproductive mode in fish and is used fairly widely in genetic breeding. Gynogenetic offspring (2n = 100, abbreviated as GRCC) were generated through the distant hybridization of Carassius auratus red var. (2n = 100, RCC) (♀) × Megalobrama amblycephala (2n = 48, BSB) (♂), in which male and female individual both had normal gonadal development. To better understand genomic and epigenetic consequences of GRCC, fluorescence in situ hybridization, amplified fragment length polymorphism, and methylation-sensitive amplification polymorphism analysis were performed on GRCC and RCC. GRCC possess two sets of RCC-derived chromosomes and one to three microchromosomes, in which 30.44% of bands inherit these patterns from red crucian carp and blunt snout bream, and 24.12% of novel bands were found by amplified fragment length polymorphism analysis. In terms of methylation, the DNA methylation level of GRCC was lower than that of their parents, and 45.29% of methylation patterns in GRCC were altered compared with their parents. GRCC show a special genetic composition in the genome, in which genome-wide changes and the adjustment of DNA methylation levels and patterns occurred. The result revealed that genetic and epigenetic changes were rapidly triggered in gynogenetic fish that were derived from distant hybridization, showing a special genetic composition in the genome. This study provides new insights into fish genetic breeding and the evolutionary patterns of the vertebrate genome.

Comparative analyses of the Sox9a-Amh-Cyp19a1a regulatory Cascade in Autotetraploid fish and its diploid parent

BackgroundAutotetraploid Carassius auratus (4nRCC, 4n = 200, RRRR) was derived from the whole genome duplication of diploid red crucian carp (Carassius auratus red var.) (2nRCC, 2n = 100, RR). To investigate the genetic effects of tetraploidization, we analyzed DNA variation, epigenetic modification and gene expression changes in the Sox9a-Amh-Cyp19a1a regulatory cascade between 4nRCC and 2nRCC.ResultsWe found that the Sox9a gene contained two variants in 2nRCC and four variants in 4nRCC. Compared with that in 2nRCC, DNA methylation in the promoter regions of the Amh and Cyp19a1a genes in 4nRCC was altered by single nucleotide polymorphism (SNP) mutations, which resulted in the insertions and deletions of CpG sites, and the methylation levels of the Sox9a, Amh and Cyp19a1a genes increased after tetraploidization. The gene expression level of the Sox9a-Amh-Cyp19a1a regulatory cascade was downregulated in 4nRCC compared with that in 2nRCC.ConclusionThe above results demonstrate that tetraploidization leads to significant changes in the genome, epigenetic modification and gene expression in the Sox9a-Amh-Cyp19a1a regulatory cascade; these findings increase the extant knowledge regarding the effects of polyploidization.

Hox genes reveal variations in the genomic DNA of allotetraploid hybrids derived from Carassius auratus red var. (female) \xd7 Cyprinus carpio L. (male)

BackgroundHox transcription factors are master regulators of animal development. Although highly conserved, they can contribute to the formation of novel biological characteristics when modified, such as during the generation of hybrid species, thus potentially serving as species-specific molecular markers. Here, we systematically studied the evolution of genomic sequences of Hox loci in an artificial allotetraploid lineage (4nAT, 4n = 200) derived from a red crucian carp (♀, RCC, 2n = 100) × common carp (♂, CC, 2n = 100) cross and its parents (RCC and CC).ResultsPCR amplification yielded 23 distinct Hox gene fragments from 160 clones in 4nAT, 22 fragments from 90 clones in RCC, and 19 fragments from 90 clones in CC. Sequence alignment of the HoxA3a and HoxC10a genes indicated both the inheritance and loss of paternal genomic DNA in 4nAT. The HoxA5a gene from 4nAT consisted of two subtypes from RCC and two subtypes from CC, indicating that homologous recombination occurred in the 4nAT hybrid genome. Moreover, 4nAT carried genomic pseudogenization in the HoxA10b and HoxC13a loci. Interestingly, a new type of HoxC9a gene was found in 4nAT as a hybrid sequence of CC and RCC by recombination in the intronic region.ConclusionThe results revealed the influence of Hox genes during polyploidization in hybrid fish. The data provided insight into the evolution of vertebrate genomes and might be benefit for artificial breeding programs.

The Hybrid Genome of a New Goldfish-Like Fish Lineage Provides Insights Into the Origin of the Goldfish.

Distant hybridization leads to obvious changes in genotypes and phenotypes, giving rise to species with novel capabilities. However, the fusion of distinct genomes also polymerizes the DNA or gene variations that occur during the course of evolution. Knowledge of the early stages of post-hybridization evolution is particularly important. Here, we investigated the full-length (FL) transcriptomes and the sequences resulting from the genome resequencing of the red crucian carp-like homodiploid fish (RCC-L) and goldfish-like homodiploid fish (GF-L) derived from the interspecific hybridization of koi carp (KOC) and blunt snout bream (BSB) to provide molecular evidence for the hybrid origin of the goldfish (GF). We compared the orthologous genes in the transcriptomes of RCC-L and GF-L with those of KOC and BSB. We also mapped the orthologous genes to the common carp (CC) and BSB genomes and classified them into eight gene patterns in three categories (chimaera, mutant, and biparental origin genes). The results showed that 48.20% and 46.50% of the genes were chimaera and that 3.70% and 8.30% of the genes were mutations of orthologous genes in RCC-L and GF-L, respectively. In RCC-L and GF-L, 63.70% and 68.20% of the genetic materials were from KOC, and 12.30% and 11.90% of the genetic materials were from BSB. The sequences from the genome resequencing of RCC-L and GF-L were mapped to the genome sequences of CC and BSB, revealing that the similarities of both RCC-L and GF-L to the CC genome (92.52%, 90.18%) were obviously higher than to the BSB genome (50.33%, 49.18%), supporting the suggestion that the genomes of both RCC-L and GF-L were mainly inherited from KOC but had some DNA fragments from BSB. Overall, our results provide molecular biological evidence for the hybrid origin of red crucian carp (RCC) and GF.

Algal Growth Enhances Light-Mediated Limitation of Bacterial Nitrification in an Aquaculture System

The accumulation of ammonia and nitrite in natural water and aquaculture systems would suppresses the immune system of aquatic animal and reduces the fish growth. Nitrifying bacteria have been widely used to reduce the accumulation of ammonia and nitrite in aquaculture systems, but are still ineffective in many cases. An aquaculture model system consisting of red crucian carp, algae, nitrifying bacteria, and pond water from a natural fish culture was established to explore the limitation of algae and light to nitrifying bacteria content and bacterial nitrification in the presence of a predator. The concentrations of nitrifying bacteria and bacterial nitrification in the group containing algae and light were significantly limited, and addition of nitrifying bacteria in algae groups had little effect. In algae-free groups, the concentrations of ammonia and nitrite were decreased by nitrifying bacteria, and the potential ammonia oxidization rate was also increased. Our findings reveal that the combined effects of algae growth and light exposure are responsible for the observed ineffectiveness of nitrifying bacteria in natural aquaculture environments.

Differential expression of activin βA and βB genes in female allotriploid and diploid red crucian carp Carassius auratus red var.

activin βA and βB from diploid and allotriploid crucian carp were cloned.The differential expression of activin βA and βB genes in female allotriploid and diploid red crucian carp Carassius auratus red var. were studied and found to be expressed in all the tested tissues; particularly, the expression of activin βA and βB was elevated in the ovaries of allotriploids and differential expression in pituitaries during the non-breeding season and the breeding season period. The immunohistochemistry indicated that the abnormal triploid ovaries were dominated by small oogonium-like cells with dense signals and that the elevated expression of activin βA and βB in the ovaries of allotriploids may be related to allotriploid sterility.

Unreduced diploid sperm from diploid hybrids and formation of a new type of tetraploid hybrid

The production of unreduced gametes is an important resource for the formation of polyploids. Here, we obtained a second generation of diploid hybrids (2nKCF2) derived from koi carp (KOC; 2n = 100; ♀) × colored crucian carp (CCC; 2n = 100; ♂) with the unusual reproductive trait of producing unreduced diploid sperm. Compared with KOC and CCC, 2nKCF2 had unique morphological characteristics, including a gray and golden head, golden-brown scaly outer edge, and smooth golden-yellow back. FISH and 5S rDNA analysis revealed that 2nKCF2 not only inherited genomic DNA from their parents but also lost some parental DNA fragments. Chromosomal spreads of germ cells provided direct cytological evidence for the production of unreduced diploid sperm by male 2nKCF2. Electron microscopy results showed that the heads of the diploid sperm generated by 2nKCF2 were twice as large as those of the haploid sperm generated by CCC (P < 0.05), confirming that the sperm of 2nKCF2 were diploid. A new type of tetraploid hybrid (4n = 200) was produced by crossing 2nKCF2 males with improved female allotetraploid hybrids of red crucian carp (♀) × common carp (♂). This study provides insights into reproductive biology and aquaculture.