Xiaoyun Zhou

Sinotaia angularis is a freshwater viviparid snail with limited genomic resources. Here, we report a chromosome-level reference genome generated using PacBio HiFi sequencing and Hi-C scaffolding, with mitochondrial marker -based screening (16S rRNA and COI (cox1)) and phylogenetic analysis supporting the taxonomic assignment. A total of 74.08 Gb of HiFi reads were obtained, providing approximately 65.7-fold genome coverage. The final assembly spans 1.127 Gb, with a scaffold N50 of 141.87 Mb and a GC content of 34.47%. Hi-C scaffolding anchored 978.89 Mb (86.85% of the assembly) onto eight chromosome-level scaffolds. BUSCO analysis using the mollusca_odb10 dataset recovered 86.9% complete orthologs from the genome assembly. Repeat annotation identified 378.75 Mb of repetitive sequences (33.60% of the genome), with unclassified repeats and LTR elements as the dominant components. Gene annotation predicted 22,232 protein-coding genes, 209 tRNAs, 72 rRNAs, 88 snRNAs, and 10 snoRNAs. Functional annotation assigned database support to 155,611 predicted proteins or isoforms, corresponding to 98% of the total protein set. CAZy annotation identified 5371 carbohydrate-active enzyme entries, suggesting broad carbohydrate-processing potential. This genome provides a reference resource for comparative genomics, chromosome evolution, repeat dynamics, gene-family evolution, and freshwater adaptation studies in Viviparidae.

The commercial viability of B. purificata relies on efficient depuration. While P. pulchellum extract dramatically accelerates this process, its physiological and molecular mechanisms remain unknown. To address this, we integrated behavioural, histological, ultrastructural, and transcriptomic (RNA-seq) analyses to characterize the acute stress and subsequent recovery phases of B. purificata exposed to the botanical extract. Extract exposure induced severe neuromuscular hyperextension response, with histological analysis revealing acute interstitial oedema and neuronal chromatolysis. This structural damage caused sustained cephalopodium extension and muscle fibre uncoupling. Transcriptomic profiling linked this neuromuscular dysfunction to conserved calcium-dependent adrenergic signalling modules and profound endogenous neuroendocrine disruption. The extract also induced cellular stress, downregulating the apoptosis inhibitor BIRC7 and eliciting transcriptomic signatures consistent with a DNA damage response. Crucially, during the short-term recovery phase, surviving tissues mounted a robust transcriptomic repair response. The snails systemically suppressed the cell cycle via MCM2-6 downregulation while upregulating GADD45 and RAD54B, suggesting a prioritization of genomic repair alongside partial morphological reorganization. Ultimately, P. pulchellum accelerates depuration via acute phytochemical stress and neuromuscular dysfunction. Importantly, this stress is accompanied by a highly coordinated transcriptomic repair response and partial short-term restoration, providing foundational molecular insights essential for evaluating and optimizing botanical depuration protocols.

The freshwater snail Bellamya purificata is both ecologically and economically significant, exhibiting notable sexual dimorphism in growth and nutritional traits that underscore the importance of breeding of monosex stocks. However, the genetic basis of sex determination remains unclear. Herein, genome-wide association studies (GWASs) combined with transcriptomic analysis were conducted to identify sex-linked markers and candidate genes for this species. GWAS generated 571 significantly sex-associated SNPs and 1853 InDels, corresponding to 44 candidate genes. Multiple significant SNP peaks were detected on chromosomes 1 and 2, with mrc2 and mis18bp1 as key candidate genes. A sex-linked InDel marker located within mis18bp1 can distinguish males and females cost-effectively. Genotype analysis of the sex-associated loci revealed that most females were homozygous while males were heterozygous, suggesting that B. purificata has a primarily XX/XY sex determination system. Comparative gonadal transcriptome analyses identified 2996 female-biased and 4281 male-biased genes. Among them, sry, sox8, dmrt1 and dmrt2 may be critical in male sex differentiation, while β-catenin, foxl2, esr1 and nr5a2 may be important in female sex differentiation. Integration of GWAS and transcriptomic data highlighted four pronounced sex-associated candidate genes, including mis18bp1, rnf216, tbx1 and mrc2. These results provide a valuable foundation for elucidating the genetic mechanisms underlying sex determination and for the development of monosex stocks in B. purificata.

Loach (Misgurnus anguillicaudatus) is a small benthic fish favored by consumers in East Asia. Female loaches exhibit greater production value due to their larger size and higher meat yield. Therefore, controlling the sex of loach should be studied to implement an all-female breeding strategy. This study identified sex-related SNP markers and candidate genes in loach through GWAS. Genotyping male and female populations with WGRS revealed 84 SNPs labeled as associated with sex. Distinct SNP peaks appeared on Chr6 and Chr3, with over half of the sex-associated SNPs located on Chr6. Within these sex-associated regions, 15 key candidate genes related to sex and reproduction were identified, including hemgn and foxl2a. Among these genes, pik3cb on Chr3 and hhip, cntln, and pard3ba on Chr6 have multiple sex-associated haplotypes that can be utilized for subsequent monosex breeding. Notably, a highly male-linked SNP marker was identified on pard3ba, whose genotype is consistent with the male heterogamete (XX/XY). Through gene enrichment analysis and expression validation, signaling pathways such as TGF-β, FoxO, and mTOR were identified as being involved in sex regulation. This study provides molecular markers for sex-controlled breeding in loach, facilitating in-depth research into the mechanisms of sex regulation in loach.

Animal mitochondrial DNA is usually considered to comply with strict maternal inheritance, and only one mitochondrial DNA haplotype exists in an individual. However, mitochondrial heteroplasmy, the occurrence of more than one mitochondrial haplotype, has recently been reported in some animals, such as mice, mussels, and birds. This study conducted extensive field surveys to obtain representative samples to investigate the existence of paternal inheritance of mitochondrial DNA (mtDNA) in natural fish populations. Evidence of paternal mitochondrial DNA leakage of P. dabryanus was discovered using high-throughput sequencing and bioinformatics methods. Two distinct mitochondrial haplotypes (16,569 bp for haplotype I and 16,646 bp for haplotype II) were observed, differing by 18.83% in nucleotide sequence. Phylogenetic analysis suggests divergence between these haplotypes and potential interspecific hybridization with M. anguillicaudatus, leading to paternal leakage. In natural populations of P. dabryanus along the Yangtze River, both haplotypes are present, with Type I being dominant (75% copy number). Expression analysis shows that Type I has higher expression levels of ND3 and ND6 genes compared to Type II, suggesting Type I’s primary role. This discovery of a species with two mitochondrial types provides a model for studying paternal leakage heterogeneity and insights into mitochondrial genome evolution and inheritance.

The diploids and tetraploids of cyprinid loach Misgurnus anguillicaudatus coexist in many natural habits. The tetraploids generally exhibit superior growth performance and delayed gonadal development compared with diploids. To investigate the regulation mechanism of growth superiority and ovarian development in tetraploids, we first conducted a long-term culture experiment and histological observation. The tetraploids exhibited a higher growth performance and delayed ovarian development. Meanwhile, at a genetic level, an average of 6891 differentially expressed genes (DEGs) between diploids and tetraploids were identified from the brain, pituitary, liver, gonad and muscle using the RNA-seq method. Among these DEGs, as expected, some candidate genes, including IGF family genes, somatostatin, leptin, cyp19a1b, gthα, lhβ and fshβ, were detected, which play critical roles in the regulation of growth and gonad development of fish. In particular, the genes related to GH/IGF axis and growth factors, signal transduction, gonadal hormone and appetite were significantly increased in tetraploids. The clustering analyses of the key candidate genes showed that most key genes were up-regulated in the pituitary and gonad of tetraploids instead of other tissues. The dynamics of these key genes provide valuable genetic evidence for clarifying the growth superiority and delayed gonadal development of tetraploids. Moreover, this study also provides some clues for unveiling the genetic superiority of polyploidy species in other phenotypes.

Triplophysa tenuis is an important indigenous fish in the Xinjiang Tarim River. In this study, we collected 120 T. tenuis individuals from 8 T. tenuis populations in the Tarim River. Through genotyping-by-sequencing (GBS), a total of 582,678,756 clean reads were generated for all the genotypes, and after quality filtering, 595,379 SNPs were obtained for the population genetic analyses. Multiple genetic parameters showed that the 8 T. tenuis populations had high genetic diversity. Phylogenetic tree analysis indicated that all T. tenuis individuals were divided into five branches, the individuals from the north of Tarim River were grouped into cluster 1 (SF and WS) and cluster 3 (DWQ, TKX, and KZE), while the AETS, WLWT and LF individuals from the south of Tarim River were clustered into cluster 2. The result was consistent with the admixture analysis, which supported that the 8 T. tenuis populations were clustered into three subgroups. Furthermore, the pairwise F_ST values and genetic distance indicated that there was a large genetic differentiation between WS and other T. tenuis populations. Collectively, this study provides valuable genome-wide data for the conservation of natural T. tenuis populations in the Tarim River.

Sexual and polyploidy size dimorphisms are widespread phenomena in fish, but the molecular mechanisms remain unclear. Loach (Misgurnus anguillicaudatus) displays both sexual and polyploid growth dimorphism phenomena, and are therefore ideal models to study these two phenomena. In this study, RNA-seq was used for the first time to explore the differentially expressed genes (DEGs) between both sexes of diploid and tetraploid loaches in four tissues (brain, gonad, liver, and muscle). Results showed that 21,003, 17, and 1 DEGs were identified in gonad, liver, and muscle tissues, respectively, between females and males in both diploids and tetraploids. Regarding the ploidy levels, 4956, 1496, 2187, and 1726 DEGs were identified in the brain, gonad, liver, and muscle tissues, respectively, between tetraploids and diploids of the same sex. When both sexual and polyploid size dimorphisms were considered simultaneously in the four tissues, only 424 DEGs were found in the gonads, indicating that these gonadal DEGs may play an important regulatory role in regulating sexual and polyploid size dimorphisms. Regardless of the sex or ploidy comparison, the significant DEGs involved in glycolysis/gluconeogenesis and oxidative phosphorylation pathways were upregulated in faster-growing individuals, while steroid hormone biosynthesis-related genes and fatty acid degradation and elongation-related genes were downregulated. This suggests that fast-growing loaches (tetraploids, females) have higher energy metabolism levels and lower steroid hormone synthesis and fatty acid degradation abilities than slow-growing loaches (diploids, males). Our findings provide an archive for future systematic research on fish sexual and polyploid dimorphisms.

The distribution of Triplophysa yarkandensis is restricted to Xinjiang’s Tarim River basin. We collected 119 T. yarkandensis samples from nine geographic populations in the Tarim River basin and utilized the RAD-seq method for SNP genotyping. In this study, a total of 164.81 Gb bases were generated with the Illumina platform, and 129,873 candidate SNPs were obtained with the Stacks pipeline for population genetic analyses. High levels of genetic diversity were detected among nine populations. The AMOVA results showed that the majority of genetic variations originated from among populations (F_ST = 0.67), and the pairwise F_ST values ranged from 0.4579 to 0.8736, indicating high levels of genetic differentiation among these populations. The discriminate analysis of principal components (DAPCs) and neighbor joining (NJ) tree revealed that the nine populations could be separated into two clusters (i.e., south and north populations), and modest genetic differentiation between south and north populations was observed, while the individuals from several populations were not clustered together by geographical location. The evidence of two genetic boundaries between south and north populations (except TTM) was supported by barrier analysis. The Bayesian skyline plotting indicated that T. yarkandensis populations in the Tarim River basin had not experienced genetic bottlenecks, and the effective population size remained stable. This study first clarified the genetic diversity and differentiation of T. yarkandensis populations in the Tarim River basin, and it provided valuable molecular data for conservation and management of natural populations.