Comparative analysis of intestinal structure, enzyme activity, intestinal microbiota and gene expression in different segments of pufferfish (Takifugu Obscurus).

The structure of fish intestines does not have a clear regional division, while the function of the intestines may be related to their structure. Therefore, in this study, the delimitation of intestinal segments in pufferfish (Takifugu obscurus) was achieved by morphological analysis. Subsequently, enzyme activity, intestinal microbiota, and gene expression were examined to compare the differences among the pufferfish various segments. According to four morphological parameters: height of mucosa folds (HF), width of mucosa folds (WF), thickness of muscularis (TM), and cross-sectional area (CSA), the pufferfish's intestine was divided into anterior intestine (AI), middle intestine (MI), and posterior intestine (PI). The activity levels of amylase, lipase, and trypsin in the AI and MI were significantly higher than these in the PI. According to the analysis of 16S rDNA, the dominant microbiota at the phylum level in the different segments were Epsilonbacteraeota, Spirochaetes, and Proteobacteria. At the genus level, there were variations observed in the relative abundance of Brevinema, Mycobacterium, Bradyrhizobium, and Microvirga. α diversity analysis revealed that the richness indexes (Ace and Chao1) were the lowest in the MI, while β diversity analysis revealed significant difference in intestinal microbial community composition among the three intestinal segments. Furthermore, RNA-Seq was used to identify differential expression genes (DEGs) and biological pathways among the different intestinal segments. The DEGs between the AI and MI were enriched in pancreatic secretion and protein digestion and absorption, those between AI and PI were involved in ascorbate and aldarate metabolism and glutathione metabolism, and those between MI and PI were involved in steroid biosynthesis, fat digestion and absorption, vitamin digestion and absorption, and glycine, serine and threonine metabolism. In conclusion, the presented results compare and analyze the differences in various intestinal segments of pufferfish, which will be conductive to future exploration of the functions of these different segments.