High temperatures are considered one of the most significant limitations to subtropical fishery production. Largemouth bass (Micropterus salmoides) is an economically important freshwater species grown in subtropical areas, which are extremely sensitive to heat stress (HS). However, comprehensive transcriptomic data for the livers of largemouth bass in response to HS are still lacking. In this study, a comparative transcriptomic analysis was performed to investigate the gene expression profiles of the livers of largemouth bass under HS treatment. As a result, 6114 significantly differentially expressed genes (DEGs), which included 2645 up-regulated and 3469 down-regulated genes, were identified in response to HS. Bioinformatics analyses demonstrated that the \'ECM-receptor interaction\' pathway was one of the most dramatically changed pathways in response to HS, and eight DEGs assigned to this pathway were taken as hub genes. Furthermore, the expression of these eight hub genes was determined by quantitative reverse transcription PCR, and all of them showed a significant change at the transcriptional level, suggesting a crucial role of the \'ECM-receptor interaction\' pathway in the response of largemouth bass to HS. These findings may improve our understanding of the molecular mechanisms underlying the response of largemouth bass to HS.

This study was conducted to ascertain the effect of dietary Zn on growth and health status of juvenile largemouth bass (Micropterus salmoides). Six experimental diets with Zn level of 50.17, 56.74, 73.34, 86.03, 123.94, and 209.20 mg/kg, respectively were compounded using complex amino acid-chelated zinc, and were fed to juvenile fish (5.50 ± 0.10 g) for 70 d. The specific growth rate (SGR) varied with dietary Zn level in a quadratic model and peaked at the 73.34 mg/kg group, while the feeding rate exhibited an opposite trend (P < 0.05). The condition factor, hepatosomatic index and mesenteric fat index all exhibited a tendency similar with SGR (P < 0.05). Dietary Zn level affected serum total proteins, urea, triglycerides, and glucose (P < 0.05). Serum Zn and copper levels linearly increased with dietary Zn level, while serum iron and manganese showed the opposite trend. Serum superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) increased with dietary Zn level and reached a plateau at 86.03 mg/kg. Serum complement component 3 (C3), IgM, and lysozyme also were enhanced by 73.34 mg/kg Zn. Body protein content increased with zinc level up to 73.34 mg/kg, and then remained steadily. As dietary Zn level increased, hepatic lipid level increased and then reached a plateau at 86.03 mg/kg group, while glycogen increased linearly. Moreover, gene expression related to lipid and glycogen metabolism from liver transcriptome further explained the liver lipid and glycogen variations. To conclude, a dietary Zn requirement of 76.99 mg/kg was suggested for juvenile largemouth bass to improve growth, antioxidant capacity, and immune status.

The present study was conducted to investigate the regulatory mechanism of liver injury in largemouth bass Micropterus salmoides (LMB) fed low protein high starch diets. Two isolipidic and isoenergetic diets were formulated with different protein and starch ratios, being named as diets P49S9 (48.8 % protein and 9.06 % starch) and P42S18 (42.4 % protein and 18.2 % starch). Each diet was fed to triplicate replicates of LMB (initial body weight, 4.65 ± 0.01 g) juveniles. Fish were fed to visual satiation for 8 weeks. The results indicated that though the P42S18 fish up-regulated the feeding ratio to meet their protein requirements, feeding efficiency ratio and growth performance were impaired in treatment P42S18 as compared to treatment P49S9. Periodic acid-Schiff (PAS) staining showed glycogen accumulated in the liver of LMB fed low protein high starch diets, and the reason should be attributed to down-regulated expression of the glycogenolytic glycogen debranching enzyme. Lower liver lipid level was associated with feeding low protein high starch diets in LMB, which should be resulted from the changes in hepatic glycerolipid metabolism regulated by lipoprotein lipase (representative of triglyceride synthesis, up-regulated) and diacylglycerol acyltransferase (representative of triglyceride breakdown, down-regulated). Though fasting plasma glucose level was comparable, treatment P42S18 performed inferior glucose tolerance to treatment P49S9. Hematoxylin-eosin (HE) and TdT-mediated dUTP Nick-End Labeling (TUNEL) staining suggested that feeding low protein high starch diets induced disruption of structural integrity, inflammation and apoptosis in the hepatocytes of LMB. As expected, KEGG pathways analysis indicated that many of the up-regulated differentially expressed genes were enriched in AGE (advanced glycation end product)/RAGE (receptor for AGE), Toll-like receptor and apoptosis signaling pathways. Our transcriptome data revealed that feeding low protein high starch diets might promote the accumulation of AGEs in LMB, which bound to RAGE and subsequently induced PI3K/Akt signal pathway. The activation of Akt induced NF-κB translocation into the nucleus thus releasing proinflammatory factors including tumor necrosis factor-α (TNF-α) and interleukin-8. The release of these inflammatory factors concomitantly induced T cell stimulation and natural killer cells chemotactic effects through Toll-like receptor signaling pathway. Besides mediating inflammation and immune response, TNF-α signal transduction participated in mediating apoptosis through the receptor of TNF (TNF-R1) pathway by up-regulating the expression of caspase 8 and cytochrome c. In conclusion, our results demonstrated that feeding low protein and high starch diets induced hepatocytes inflammation and apoptosis in LMB through the PI3K/Akt/NF-κB signaling pathway.

The present study was conducted to investigate the effects of bile acids (BAs) on the growth, liver function and immunity of the largemouth bass fed high-starch diet. The experiment set three isonitrogenous and isoenergetic semi-purified diets, LS: low-starch diet (5%), HS: high-starch diet (19%) and SB: high-starch diet with BAs (350 mg/kg diet). An 8-week feeding trial was conducted in largemouth bass of initial weight 23.69 ± 0.13 g. The results indicated that the weight gain (WG) and protein efficiency ratio (PER) of fish fed LS and SB were significantly higher than HS treatment. The superoxide dismutase (SOD) and catalase (CAT) activities of SB group were significantly increased, while malondialdehyde (MDA) content significantly reduced in liver compared with HS group. The activities of alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) and glucose contents in plasma of SB group were significantly lower than HS treatment, whereas the content of triglyceride (TG) and total cholesterol (TC) in plasma were significantly higher than HS treatment. Additionally, the plasma immunoglobulin count, lysozyme activity and the blood leukocyte count (WBC) in SB group were significantly higher than HS group. The results of paraffin section of liver showed the histopathological alterations were significantly reduced in the SB group compared to HS group. All in all, this study revealed that bile acids supplement could significantly improve growth performance, enhance liver function and immune ability, and alleviate stress responses of M. salmoides fed high-starch diet.

The present study was conducted to investigate the effects of high dietary lipid levels on growth, metabolism, antioxidant capacity, and immune responses of largemouth bass. Fish (initial body weight 13.38 ± 0.11 g) were fed three isonitrogenous semi-purified diets containing 5%, 10%, and 20% lipid, respectively. The results indicated that fish fed 10% lipid diet showed significantly better final body weight, specific growth rate (SGR), protein efficiency ratio (PER), and feed conversion ratio (FCR) compared with that fed 5% lipid diet. Meanwhile, fish fed 20% lipid diet had a significantly higher viscera ratio (VR), hepatosomatic index (HSI), intraperitoneal fat ratio (IPF), and liver lipid content than those fed the other diets. Higher alanine aminotransferase (ALT) and aspartate transaminase (AST) activities, total cholesterol (TC), triglyceride (TG), free fatty acids (FFA), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C) contents, and LDL-C/HDL-C value in plasma were recorded in fish fed 20% lipid diet, while higher insulin contents were obtained in fish fed 5% lipid diet. In addition, the highest carnitine palmitoyltransferase I (CPT1), AMP-activated protein kinase (AMPK), fructose-1,6-bisphosphatase (FBPase), and phosphoenolpyruvate carboxykinase (PEPCK) activities in the liver were also observed in fish fed 20% lipid diet. However, fish fed 20% lipid diet had a significantly lower superoxide dismutase (SOD) and catalase (CAT) activities and higher MDA contents in liver than those fed the other diets. The higher nitric oxide (NO) contents and inducible nitric oxide synthase (iNOS) activity in liver were recorded in fish fed 10% lipid diet. Moreover, the alkaline phosphatase (ALP), inducible nitric oxide synthase (iNOS) and lysozyme activities, and nitric oxide (NO) contents in plasma were higher in fish fed the 10% diets than the other groups. In conclusion, high dietary lipid levels could suppress growth performance and liver anti-oxidative capacity, and reduce immune responses of largemouth bass.

A feeding trial was conducted to evaluate the effect of linseed oil (LO) on growth, plasma biochemistry, hepatic metabolism enzymes, and antioxidant capacity of juvenile largemouth bass, Micropterus salmoides. Four isonitrogenous (crude protein, 45%) and isoenergetic (gross energy, 18 MJ/kg) diets were formulated by replacing 0 (the control), 33.3%, 66.7%, and 100% of fish oil with linseed oil. Each diet was fed to three replicate groups of fish (initial body weight, 22.02 ± 0.61 g) for 8 weeks. The results indicated that fish fed diet with 100% LO substitution level had lower weight gain (WG), specific growth rate (SGR), and protein efficiency ratio (PER) than the other groups (P < 0.05), while feed conversion ratio (FCR) was higher compared to the other groups (P < 0.05). Feed intake (FI) and hepatosomatic index (HSI) of 66.7% LO substitution level were significantly lower than the control groups (P < 0.05). Glycogen, lipid, and non-esterified fatty acid content in the liver decreased significantly with increasing dietary LO levels (P < 0.05). Moreover, the replacement of fish oil (FO) with LO could significantly reduce the content of triglyceride (TG) and total cholesterol (TC) and the activity of alanine amiotransferase (ALT) in plasma of M. salmoides (P < 0.05). There were significant differences in hepatic metabolism enzymes in fish fed diets with different dietary LO levels. Adenosine 5\'-monophosphate (AMP)-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor (PPAR-α) activities in liver significantly increased with increasing dietary LO level (P < 0.05). In addition, phosphoenolpyruvate carboxykinase (PEPCK) and fructose-1,6-bisphosphatase (FBPase) activities in the liver significantly increased with decreasing dietary LO level (P < 0.05). Both the lowest superoxide dismutase (SOD) and catalase (CAT) activities in the liver were recorded in the control group (P < 0.05). Moreover, nitric oxide content, glutathione peroxidase (GPx), and inducible nitric oxide synthase (iNOS) activities in the liver significantly increased with increasing dietary LO level, while malondialdehyde (MDA) content significantly reduced. These findings demonstrated that LO can improve liver function and antioxidant ability of M. salmoides. In addition, replacing partial FO with LO cannot affect growth performance, but all substitutions inhibit growth performance of M. salmoides.