Photoheterotroph improved the growth and nutrient levels of Chlorella vulgaris and the related molecular mechanism.

Microalgae are rich in fatty acids, proteins, and other nutrients, which have gained the general attention of researchers all over the world. For the development of Chlorella vulgaris in food and feed industry, this study was conducted to investigate the differences in C. vulgaris' growth and nutritional components under different culture conditions (autotrophic, heterotrophic, photoheterotrophic) and the internal factors through cell counting in combination with transcriptome and nutrient analyses. The results showed that, under the photoheterotrophic condition, Chlorella's growth and the contents of lipid and protein were significantly higher than that under the heterotrophic condition, and the moisture content was lower than that under the heterotrophic condition. The saturated fatty acid content under the photoheterotrophic condition was the lowest, while the polyunsaturated fatty acid content was significantly higher than those under the other two conditions. There were 46,583 differentially expressed genes (DEGs), including 33,039 up-regulated DEGs (70.93%) and 13,544 down-regulated DEGs (29.07%), under the photoheterotrophic condition in comparison with the autotrophic condition. The fold change between the two conditions of samples of up-regulated genes was higher than that of the down-regulated genes. The KEGG enrichment showed that the up-regulated DEGs in the photoheterotrophic condition were significantly enriched in 5 pathways, including protein processing in endoplasmic reticulum pathway, photosynthesis pathway, photosynthesis-antenna protein pathway, endocytosis pathway, and phosphonate and phosphinate metabolism pathway. DEGs related to fatty acid metabolic pathways were significantly enriched in the fatty acid biosynthesis pathway and the biosynthesis of unsaturated fatty acid pathway. The qPCR analysis showed that the expression pattern of the selected genes was consistent with that of transcriptome analysis. The results of this study lay a theoretical foundation for the large-scale production of Chlorella and its application in food, feed, and biodiesel. KEY POINTS: • Nutrient levels under photoheterotrophic condition were higher than other conditions. • Six important pathways were discovered that affect changes in nutritional composition. • Explored genes encode important enzymes in the differential metabolic pathways.

Establishment and characterization of a skeletal myoblast cell line of grass carp (Ctenopharyngodon idellus).

Skeletal muscle myoblastic cell lines can provide a valuable new in vitro model for the exploration of the mechanisms that control skeletal muscle development and its associated molecular regulation. In this study, the skeletal muscle tissues of grass carp were digested with trypsin and collagenase I to obtain the primary myoblast cell culture. Myoblast cells were obtained by differential adherence purification and further analyzed by cryopreservation and resuscitation, chromosome analysis, immunohistochemistry, and immunofluorescence. A continuous grass carp myoblast cell line (named CIM) was established from grass carp (Ctenopharyngodon idellus) muscle and has been subcultured > 100 passages in a year and more. The CIM cells revived at 79.78-95.06% viability after 1-6 months of cryopreservation, and shared a population doubling time of 27.24 h. The number of modal chromosomes of CIM cells was 48, and the mitochondrial 12S rRNA sequence of the CIM cell line shared 99% identity with those of grass carp registered in GenBank. No microorganisms (bacteria, fungi, or mycoplasma) were detected during the whole study. The cell type of CIM cells was proven to be myoblast by immunohistochemistry of specific myogenic protein markers, including CD34, desmin, MyoD, and MyHC, as well as relative expression of key genes. And the myogenic rate and fusion index of this cell line after 10 days of induced differentiation were 8.96 ~ 9.42% and 3-24%, respectively. The telomerase activity and transfection efficiency of CIM cell line were 0.027 IU/mgprot and 23 ~ 24%, respectively. These results suggest that a myoblast cell line named CIM with normal biological function has been successfully established, which may provide a valuable tool for related in vitro studies.

Effects of Dietary Cottonseed Protein Concentrate Levels on Growth Performance, Health Status, Flesh Quality and Intestinal Microbiota of Grass Carp (Ctenopharyngodon idellus).

The aim of this study was to evaluate the nutritional value of cottonseed protein concentrate (CPC) as a single dietary protein source and the optimal protein level for grass carp (Ctenopharyngodon idellus). An 8-week feeding trial was conducted by feeding juvenile grass carp (initial body weight: 4.68 ± 0.01 g) with six experimental diets containing graded levels of protein provided by CPC. The results showed that the optimal CPC level (CPC4) improved the growth performance and health status of grass carp. The optimal dietary protein level was estimated to be 38.61 and 38.66% based on specific growth rate (SGR) and feed efficiency (FE), respectively. The CPC4 group significantly increased the total antioxidant capacity (T-AOC) content and glutathione peroxidase (GSH-Px) activity in the hepatopancreas (p < 0.05). In addition, the CPC4 group increased the muscle T-AOC and glutathione (GSH) content and improved muscle hardness, and the gene expression of MRFs, fgf6a, myhc-7, myhc-1, myhc-4, igf-II, and tor was upregulated while mstn gene expression was downregulated (p < 0.05). Correlation analysis revealed that the optimal dietary CPC level promoted grass carp growth, health, and flesh quality by regulating the relative abundance of intestinal microbes. Furthermore, CPC6 upregulated the ko00480 (Glutathione metabolism) and ko00620 (Pyruvate metabolism) pathways compared to CPC1 (p < 0.05), possibly indicating that low dietary CPC levels adversely affected amino acid metabolism in the intestinal microbiota of grass carp, while a high level of CPC will meet the metabolic needs of the body by increasing the utilization of energy.

The Interaction of Dried Distillers Grains With Solubles (DDGS) Type and Level on Growth Performance, Health, Texture, and Muscle-Related Gene Expression in Grass Carp (Ctenopharyngodon idellus).

The aim of this study was to estimate the possible synergetic effects of the two levels of dietary dried distillers grains with solubles (DDGS) from different sources (US-imported and native) on the growth, health status, muscle texture, and muscle growth-related gene expression of juvenile grass carp. Four treatments of fish were fed with 4 isonitrogenous diets, namely, native DDGS20, native DDGS30, US-imported DDGS20, and US-imported DDGS30 for 60 days. The US-imported DDGS30 group showed the better growth and feed efficiency. Additionally, we observed a significant increase in hepatopancreatic total antioxidant capacity (T-AOC) and superoxide dismutase (SOD) in native DDGS groups. Moreover, raw muscle collagen increases considerably in the US-imported DDGS30 compared with the native DDGS30 group. In comparison with the native DDGS groups, the US-imported DDGS groups showed significantly decrease in all textural properties and fiber density, while increased fiber diameter. Dietary native DDGS inclusion significantly showed the upregulation of myog, myhc, and fgf6a expression in muscle, while the downregulation of the expression of myod and myf5. Overall, US-imported DDGS30 had a beneficial influence on growth via regulating genes involved in myogenesis and hypertrophy, the formation of collagen, but had negative impacts on antioxidant capacity and cooked muscle texture.