Effects of Coatings on Antioxidant Enzyme Activities, Histopathology, and Transcriptome Profiles of Kidney Tissue in Larimichthys crocea

Abstract

Background: As an innovative approach to deep-sea aquaculture, fish farm vessels offer a dual benefit by alleviating the pressure on offshore fishing resources while providing an additional high-quality protein source. However, the potential impacts of vessel coatings on farmed fish remain poorly understood. Methods: In this study, to investigate the effects of vessel coatings on the large yellow croaker (Larimichthys crocea), we established four experimental groups with coating concentrations at 1-fold, 10-fold, 20-fold, and 80-fold levels. Antioxidant enzyme activities in kidney tissues were measured across all groups, while histological and transcriptome analyses were specifically conducted for the 1-fold and 80-fold concentration groups. Results: Firstly, significant alterations in antioxidant enzyme activity were observed in the 80-fold concentration group. Moreover, histological analysis demonstrated more severe pathological changes in kidney tissue at the higher concentration, including interstitial hemorrhage and tubular epithelial cell fatty degeneration. In addition, we identified 11,902 differentially expressed genes (DEGs) by high-throughput sequencing. KEGG pathway enrichment analysis revealed that the DEGs were predominantly involved in critical biological processes, including endoplasmic reticulum protein processing, oxidative phosphorylation, cytokine-cytokine receptor interactions, cell cycle regulation, DNA replication, and PPAR signaling pathways. Finally, the validation of nine selected DEGs through quantitative real-time PCR (qRT-PCR) showed significant correlation with RNA-Seq data, confirming the reliability of our transcriptome analysis. Conclusions: This study provides preliminary insights into the antioxidant stress response mechanisms of L. crocea to coating exposure and establishes a theoretical foundation for optimizing healthy fish farming practices in aquaculture vessels.

Keywords: RNA-seq; antioxidant enzyme activities; aquaculture vessel coatings; fish; histology.

Figure 1

Activity of peroxidase (POD) in the kidneys of L. crocea under (A) 1-fold, (B) 10-fold, (C) 20-fold, and (D) 80-fold coating concentration. * means p < 0.05; ** means p < 0.01.

Figure 2

Contents of malondialdehyde (MDA) in the kidneys of L. crocea under (A) 1-fold, (B) 10-fold, (C) 20-fold, and (D) 80-fold coating concentration. * means p < 0.05; ** means p < 0.01.

Figure 3

Activity of catalase (CAT) in the kidneys of L. crocea under (A) 1-fold, (B) 10-fold, (C) 20-fold, and (D) 80-fold coating concentration. * means p < 0.05; ** means p < 0.01.

Figure 4

Activity of superoxide dismutase (SOD) in the kidneys of L. crocea under (A) 1-fold, (B) 10-fold, (C) 20-fold, and (D) 80-fold coating concentration. * means p < 0.05; ** means p < 0.01.

Figure 5

Effects of coatings on kidney histopathological features of L. crocea. (A,D) Kidney tissue structures in the control group; (B,E) kidney tissue structure at 24 h and 48 h in the 1-fold concentration group; (C,F) kidney tissue structure at 24 h and 48 h in the 80-fold concentration group. GL: glomerulus; TU: renal tubule; TN: renal tubular epithelial cell necrosis; TD: fatty degeneration of renal tubular epithelial cells.

Figure 6

Principal component analysis (PCA) of kidney transcriptome at 24 h and 48 h after coating immersion.

Figure 7

Analysis of DEGs in L. crocea kidney at different time points after coating immersion displayed through volcano plot and Venn diagram. (A,B) Volcano plots of DEGs in L. crocea kidney at 24 h and 48 h in the 1-fold concentration group, respectively. (C,D) Volcano plots of DEGs in L. crocea kidney at 24 h and 48 h in the 80-fold concentration group, respectively. The x-axis represents the value of Log₂ (fold change), and the y-axis represents the value of –Log₁₀ (FDR). Red and green spots represent significantly up-regulated and down-regulated genes, respectively (|log₂ (fold change)| > 1), while blue spots represent no difference in gene expression. (E) Venn diagram.

Figure 8

Heatmap display of hierarchical clustering of all DEGs in the kidneys of L. crocea at different time points after coatings immersion. DEGs are displayed in rows, while samples are displayed in columns.

Figure 9

Classification map of secondary DEG entries. The x-axis represents the second-level GO entry, and the y-axis represents the number of DEGs in the GO entry. (A) GO analysis of up-regulated DEGs; (B) GO analysis of down-regulated DEGs.

Figure 10

Pathway analysis of DEGs in the kidneys of L. crocea between control group and coating-treated groups based on the KEGG database. The vertical coordinates indicate the KEGG channel. The horizontal coordinates indicate the Rich factor. The larger the Rich factor, the greater the degree of enrichment. The larger the point, the greater the number of differential genes enriched in the pathway. (A) Pathway analysis of up-regulated DEGs; (B) pathway analysis of down-regulated DEGs.

Figure 11

Representative pathway analysis of DEGs in L. crocea kidney between control group and coating-treated groups based on KEGG database. Red and green represent up-regulated and down-regulated genes, respectively, while yellow represent no difference in gene expression.

Figure 12

Validation of RNA-Seq results using qPCR. Comparison of DEG expression data between RNA-seq (green bars) and qRT-PCR (red bars). The x-axis presents the time points, and the y-axis presents the Log₂ Fold change in DEG expression. (A,B) Changes in gene expression in the 1-fold concentration group at 24 h and 48 h compared with the control group; (C,D) Changes in gene expression in the 80-fold concentration group at 24 h and 48 h compared with the control group.