Pathogenic infection of fishes is an important constraining factor affecting marine aquaculture. Insufficient understanding of the molecular mechanisms has affected the diagnosis and corresponding treatment. Here, we reported the dynamic changes of gene expression patterns in the Chinese tongue sole kidney at 16 h, 48 h, 72 h and 96 h after Vibrio harveyi infection. In total, 366, 214, 115 and 238 differentially expressed genes were obtained from the 16 h−vs. −C, 48 h−vs. −C, 72 h−vs. −C and 96 h−vs. −C group comparisons, respectively. KEGG enrichment analysis revealed rapid up-regulation of several immune-related pathways, including IL-17, TNF and TLR signaling pathway. More importantly, time-series analyses of transcriptome showed that immune genes were specifically up-regulated in a short period of time and then decreased. The expression levels of chemokines increased after infection and reached a peak at 16 h. Specifically, Jak-STAT signaling pathway played a crucial role in the regulation during Vibrio harveyi infection. In the later stages of infection, genes in the neuroendocrine pathway, such as glucocorticoid-related genes, were activated in the kidney, indicating a close connection between the immune system and neuroendocrine system. Our dynamic transcriptome analyses provided profound insight into the gene expression profile and investigation of immunogenetic mechanisms of Chinese tongue sole.

Spermatogenesis is a complex and continuous process of germ-cell differentiation. This complex process is regulated by many factors, of which gene regulation in spermatogenic cells plays a decisive role. Spermatogenesis has been widely studied in vertebrates, but little is known about spermatogenesis in protochordates. Here, for the first time, we performed single-cell RNA sequencing (scRNA-seq) on 6832 germ cells from the testis of adult Ciona intestinalis. We identified six germ cell populations and revealed dynamic gene expression as well as transcriptional regulation during spermatogenesis. In particular, we identified four spermatocyte subtypes and key genes involved in meiosis in C. intestinalis. There were remarkable similarities and differences in gene expression during spermatogenesis between C. intestinalis and two other vertebrates (Chinese tongue sole and human). We identified many spermatogenic-cell-specific genes with functions that need to be verified. These findings will help to further improve research on spermatogenesis in chordates.

Innate immunity represents a foundational defense strategy across bilaterians, with phagocytosis and coagulation serving as its central effector mechanisms. However, it remains uncertain whether these mechanisms evolved from conservative ancestral regulatory modules or emerged through lineage-specific adaptations. This ambiguity currently impedes a deeper understanding of immune system evolution. We constructed a cross-phylum single-cell atlas of circulating immune cells from nine bilaterian species. Our comparative analysis revealed a pattern of evolutionary tinkering, wherein core functional modules followed largely independent evolutionary trajectories. The specific phagocytic-like cells (PLCs) identified in invertebrates share a core gene regulatory network orchestrated by the MiT/TFE transcription factor family with vertebrate myeloid cells, indicating deep homology. Notably, we identify and characterize coagulation effector cells in invertebrates for the first time. These clot-associated hemocytes demonstrate transcriptome-level convergence in the absence of a conserved regulatory framework, achieving similar functional states through lineage-specific genetic pathways. Our findings highlight the distinct evolutionary trajectories of innate immune cells, distinguishing the ancient, hardwired regulatory program of phagocytosis from the convergent, adaptive nature of coagulation. This study offers a unified single-cell perspective on the assembly and diversification of the bilaterian immune system.