Zhuangwen Mao

The majority of Aeromonas strains are opportunistic pathogens for both humans and animals, causing a variety of diseases and posing a considerable risk to their health. In the current study, triploid cyprinid fish (TCF) were infected with a novel pathogenic Aeromonas sp. AS1-4 for pathological analysis. TCFs receiving Aeromonas sp. AS1-4 challenge exhibited oxidative damage in the liver and spleen, along with significant changes in immune-related gene expressions. Metabolomics assay indicated that strain AS1-4 challenge may exhibit a significant impact on metabolic processes of amino acids, with methylsuccinic acid (MSA) identified as vital biomarker. Following that, three potential probiotics designated Enterobacter strains fkY27-2, fkY84-1 and fkY84-4 were isolated from the intestine of TCFs, displaying excellent safety characteristics. In addition, intestinal Enterobacter strains exhibited multiple probiotic traits, including high degrees of hydrophobic activity, aggregation performance, biofilm-forming activity (BFA) and nutrient decomposing ability. Moreover, these probiotic isolates markedly coaggregated with Aeromonas sp. AS1-4 and Edwardsiella tarda 1l-4 and then suppressed their pathogenic biofilm-forming abilities, along with possessing robust antioxidant potential against various free radicals. These findings may provide valuable insights into metabolic response in fish post infection and health management in aquaculture.

A 56-day feeding trial was conducted to evaluate the effect of different dietary protein sources on the growth performance, muscle flavor substances, and quality structure of Triploid Crucian Carp. Three isonitrogenous (32.00%), isolipidic (8.00%), and isoenergetic (18.00 MJ kg⁻¹) practical diets were formulated. These diets consisted of fishmeal as the animal-derived protein source (AP), a combination of soybean meal and rapeseed meal as the plant-derived protein source (PP), and a mixture of fishmeal, soybean meal, and rapeseed meal as the mixed protein source (MP). Each diet was randomly assigned to triplicate tanks of fish and each tank was stocked with 25 fish (11.5 ± 0.4 g). The fish were fed until apparent satiation twice a day. The results showed a significant enhancement in the growth performance of Triploid Crucian Carp in the AP group compared with both the MP and PP groups (p < 0.05). Dietary plant derived protein can remarkably reduce the crude lipid content and increase the moisture content of the whole body and the dorsal muscle (p < 0.05). The antioxidant ability of fish in the PP group and MP group was better than that in the AP group (p < 0.05). Regarding free amino acids composition of muscle, the contents of glycine, methionine, and lysine were significantly enhanced in the AP group, while the histidine content was significantly increased in the PP group (p < 0.05). In terms of texture, dietary plant protein significantly improved the muscular hardness, gumminess, and chewiness. The activities of intestinal trypsin and alkaline phosphatase (ALP) of fish in the PP group were significantly higher than that in the AP group (p < 0.05). The expression levels of hepatopancreas TOR and IGF1 genes in the PP group were significantly higher than that in the MP group (p < 0.05). The present results indicate dietary fishmeal significantly improved the growth performance and muscular flavor glycine content, while dietary plant-based protein increased crude protein content, antioxidant ability, and muscular texture performance of Triploid Crucian Carp.

Kidney-type glutaminase, encoded by the gls1 gene, plays a critical role in glutamate production and improvement of meat flavor. In this study, a gls1 gene encoding 595 amino acids was cloned from triploid crucian carp (Carassius auratus) (TCC) and showed a high similarity with the gls1 gene found in Cyprinus carpio, Sinocyclocheilus rhinocerous and Puntigrus tetrazona. Comparing the abundance of gls1 in different tissues, we found its expression level in the brain and liver were significantly higher than that in heart, gut, kidney, spleen and muscle. gls1 expression in the brain reached the highest value. In addition, the expression levels of gls1 also appeared different in diurnal variation, with the highest expression seen at 9:00, while it was low at 3:00, 6:00, 15:00 and 24:00. Furthermore, dietary regulation of gls1 expression was investigated in our study. In each feeding trial, each diet was randomly assigned to triplicate tanks. Fish were fed one of the tested diets up to satiation twice daily. The results showed that gls1 expression increased in 32% protein group and decreased in 35–41% protein group. The results of different protein source experiments showed that the expression of gls1 gene in the mixed protein group (the control group) was significantly higher than that in the fish meal and soybean meal groups. Glutamate treatment revealed that appropriate concentrations (0.10 mg/mL in vivo and 2.00% in vitro) of glutamate remarkably improved the expression of gls1. Besides, diets supplemented with 0.80–1.60% lysine-glutamate dipeptide exhibited a down regulatory impact on gls1 expression. In conclusion, this study demonstrated that the expression of gls1 in TCC was increased by 32% protein diet, mixed protein source diet and diet with 2.00% glutamate concentration, while decreased by 0.80–1.60% lysine-glutamate dipeptide. The findings of this study provide a reference for the regulation of gls1 and have a potential application in the optimization of dietary formula in aquaculture.

Glutamine synthetase (GS) is a key enzyme that catalyzes the synthesis of glutamine from glutamate, which plays a role in the promotion of muscle cell growth and in improving the flavor of meats. In this study, a GS gene encoding 371 amino acids was cloned from triploid crucian carp and showed the highest level of similarity with the GS gene found in Cyprinus carpio. Meanwhile, GS was differentially expressed in different tissues, and its day–night expression changes showed obvious oscillation. Additionally, the effects of glutamate and glutamine on GS expression in muscle cells were investigated in vitro and in vivo. We found that its expression was obviously increased due to high levels of glutamate (2 mg/mL) but decreased by glutamine in vitro. However, it was significantly promoted by glutamate and glutamine in vivo, with an optimal concentration of 2%. Furthermore, the use of lysine–glutamate dipeptides as feed additives also had a positive influence on GS expression (the optimal concentration is 0.8%). Finally, we explored the effects of different protein levels and sources on the expression of GS, and the results demonstrated that GS had the highest expression at the 35% protein level, but no significant differences were observed in the different protein sources between the fish meal diet (FM) and the mixed diet comprising soybean meal and rapeseed meal (SM). This study sheds new light on the regulation of GS in teleost fish and provides new perceptions and strategies for the formulation of high-quality feed for triploid crucian carp.

AlaSerCys Transporter 2 (ASCT2), encoded by the SLC1A5 gene, plays an important role in the absorption of glutamine. In this study, the full-length cDNA sequence of ASCT2 was cloned from triploid crucian carp. It encodes 539 amino acid residues and a stop codon. Phylogenetic analysis revealed that the sequences of the ASCT2 ORF region in cyprinid fishes shared high sequence homology. Comparing the abundance of ASCT2 in different tissues, we found its expression level in muscle was significantly higher than that in intestine (p < 0.05). In addition, the expression levels of ASCT2 also appeared different in diurnal variation. Then we found the addition of 2.5% glutamate in a feeding diet significantly increased the expression levels of ASCT2 in intestine and muscle (p < 0.05). However, in glutamine experiments, the muscle showed the highest expression level of ASCT2 when fish were fed the diet containing 3.0% glutamine (p < 0.05). In vitro, ASCT2 was sensitive to glutamine and its expression level appeared down-regulated when the addition of glutamine was added to 0.1 mg/mL. Finally, we found that the diet with 29% protein level significantly increased the expression level of ASCT2 in intestine (p < 0.05). Nevertheless, different protein sources (fish meal and soybean meal) had no significant effect on the expression levels of ASCT2 in intestine and muscle (p > 0.05). These results provided data for the study of ASCT2 in triploid crucian carp regulated by feeding nutrition, which had a potential application in improving feed formulation in aquaculture.