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The major histocompatibility complex (MHC) genes play a key role in immune response in vertebrates. In this study, an MHC I alpha homolog gene (PfMHC Ⅰα) from pufferfish (Takifugu obscurus) was identified and its subcellular localization and expression patterns of PfMHC Ⅰα after challenge in vivo and in vitro were analysed. The open reading frame of PfMHC Ⅰα was 1,089 bp in length, encoding 362 aa. The immunofluorescence result revealed that PfMHC Ⅰα was presented on the membrane of lymphocytes. qRT‐PCR analysis indicated that PfMHC Ⅰα was expressed in all examined tissues, with the highest expression in skin, followed by the expression in gills and whole blood. After challenge of Aeromonas hydrophila or polyinosinic: polycytidylic acid (Poly I:C) in vitro, the expression levels of PfMHC Ⅰα on pufferfish kidney lymphocytes were significantly up‐regulated, with the highest expression level at 48 hr post‐challenge. After infection with A. hydrophila or Poly I:C in vivo, the expression levels of PfMHC Ⅰα in the skin, whole blood and kidneys were significantly up‐regulated. Taken together, it is speculated that PfMHC Ⅰα associates with resistance to both intracellular and extracellular antigens and plays an important role in the host response against pathogen infection in pufferfish.

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The effects of replacing fish meal (FM) with low-gossypol cottonseed meal (LCSM) on the intestinal barrier in different segments (PI, proximal intestine; MI, middle intestine; DI, distal intestine) were studied in juvenile golden pompano (Trachinotus ovatus) using 6-week feeding trials. FM of 0% (diet_1), 20% (diet_2), 40% (diet_3) and 60% (diet_4) was replaced by LCSM. The FM-based diet (diet_1) containing 25% FM was used as control group. The intestinal morphology, mRNA expression of tight junctionrelated genes (ZO-1, Occludin, Claudin-12), antioxidant enzyme-related genes (CAT, GPx, GR, CuZnSOD) and non-specific immune response-related genes (IL-8, TNFα, C4) and microbiota of different intestine segments were detected. The results showed that the number and length of microvilli and intestinal wall thickness decreased significantly in PI of diet_3 and diet_4 groups. Meanwhile, mRNA expression of Occludin, Clauding-12 and C4 was up-regulated in PI and DI of diet_3 group; the expression of GR and CuZnSOD was up-regulated in PI and MI with increasing LCSM replacement; the expression of TNFα was up-regulated in the three segments of diet_3 group. Further, 16S rRNA amplicon sequencing revealed a significant reduction in the richness and diversity of intestinal microbial in diet_3 and diet_4 groups. At the family level and genera level, the abundance of Nocardiaceae and Rhodococcus was up-regulated in the three segments of diet_3 and diet_4 groups respectively. These results suggested that the replacement of LCSM by more than 20% causes the intestinal barrier function of juvenile golden pompano to be affected. These provide a reference for sustainable aquaculture.