Effect of dietary koumine on the immune and antioxidant status of carp (Cyprinus carpio) after Aeromonas hydrophila infection.

Koumine (KM) has anxiolytic, anti-inflammatory and growth-promoting effects in pigs and sheep. Based on the growth-promoting and immunological effects of koumine, the present study was conducted on Cyprinus carpio (C. carpio) with four KM concentrations: 0 mg/kg, 0.2 mg/kg, 2 mg/kg, and 20 mg/kg for 10 weeks, followed by a 1-week Aeromonas hydrophila (A. hydrophila) infection experiment. The effect of KM on the immunity of A. hydrophila infected carp was analyzed by histopathology, biochemical assay, and qRT-PCR to assess the feasibility of KM in aquaculture. The results showed that the presence of KM alleviated pathogen damage to carp tissues. At 2 mg/kg and 20 mg/kg concentrations of KM successively and significantly elevated (p < 0.05) the SOD activities in the intestinal tract, hepatopancreas and kidney of carp. The expression levels of hepatopancreatic antioxidant genes Nrf2 and IGF-1 were significantly up-regulated in the same group (p < 0.05), while the expression levels of immune genes IL-8 and IL-10 were down-regulated. In summary, KM at concentrations of 2 mg/kg and 20 mg/kg could regulate the expression of antioxidant and immune genes in various tissues in an orderly and rapid manner, and significantly improve the antioxidant and immune abilities of carp, which is conducive to the improvement of the resilience of carp.

Replacing starch with resistant starch (Laminaria japonica) improves water quality, nitrogen and phosphorus budget and microbial community in hybrid snakehead (Channa maculata ♀ × Channa argus ♂).

It is essential to increase the use of carbohydrates as an energy source and improve protein synthesis and utilization to reduce ammonia nitrogen emissions. A 60-day cultural experiment was conducted to assess the impact of resistant starch (kelp meal, Laminaria japonica) replacing starch on water quality, nitrogen and phosphorus budget and microbial community of hybrid snakehead. Approximately 1350 experimental fish (11.4 ± 0.15 g) were randomly divided into control group (C, 20% starch) and four resistant starch groups: low replacement group (LR, 15% starch), medium replacement group (MR, 10% starch), high replacement group (HR, 5% starch) and full replacement group (FR, 0% starch). The crude protein and crude fat content of hybrid snakehead fish fed with the FR diet had the most significant improvement (P < 0.05). However, resistant starch also increased the effectiveness of nitrogen and phosphorus utilization in hybrid snakeheads, which decreased the proportion of total nitrogen and total phosphorus in tail water. The minimum nitrogen and phosphorus emission rate was when the starch level was 6.1%. Denitrifying microbes including Gemmobacter, Rhodobacter, Emticicia and Bosea have become much more prevalent in group FR (P < 0.05). In general, replacing starch with resistant starch can enhance the rate at which nitrogen and phosphorus are used in feeding, lessening water pollution and altering environmental microbial composition. PRACTITIONER POINTS: Resistant starch (RS) improves whole fish nutritional content. Resistant starch improves dietary nitrogen and phosphorus utilization. Resistant starch acts as a carbon source and encourages the colonization of denitrifying bacteria in water.

Effects of dietary Gelsemium elegans alkaloids on intestinal morphology, antioxidant status, immune responses and microbiota of Megalobrama amblycephala.

Numerous plant extracts used as feed additives in aquaculture have been shown to stimulate appetite, promote growth and enhance immunostimulatory and disease resistance in cultured fish. However, there are few studies on the famous Chinese herbal medicine Gelsemium elegans, which attracts our attention. In this study, we used the Megalobrama amblycephala to investigate the effects of G. elegans alkaloids on fish intestinal health after diet supplementation with 0, 5, 10, 20 and 40 mg/kg G. elegans alkaloids for 12 weeks. We found that dietary G. elegans alkaloids at 40 mg/kg improved intestinal morphology by increasing villus length, muscle thickness and villus number in the foregut and midgut and muscle thickness in the hindgut (P < 0.05). These alkaloids also significantly improved intestinal antioxidant capabilities by increasing superoxide dismutase, catalase, total antioxidant capacity and malondialdehyde levels and up-regulated intestinal Cu/Zn-SOD and Mn-SOD (P < 0.05) at 20 and 40 mg/kg. Dietary G. elegans alkaloids improved intestinal immunity via up-regulating the pro-inflammatory cytokines IL-1ß, IL-8, TNF-α and IFN-α and down-regulating expression of the anti-inflammatory cytokines IL-10 and TGF-ß (P < 0.05) at 20 and 40 mg/kg. The expression of Toll-like receptors TRL1, 3, 4 and 7 were also up-regulated in intestine of M. amblycephala (P < 0.05). In intestinal microbiota, the abundance of Proteobacteria was increased while the Firmicutes abundance was decreased at phylum level after feeding the alkaloids (P < 0.05). The alkaloids also increased the abundance of the probiotic Rhodobacter and decreased the abundance of the pathogenic Staphylococcus at genus level (P < 0.05). In conclusion, dietary G. elegans alkaloid supplementation promoted intestine health by improving intestine morphology, immunity, antioxidant abilities and intestinal microbiota in M. amblycephala.

Effects of dietary Gelsemium elegans alkaloids on growth performance, immune responses and disease resistance of Megalobrama amblycephala.

The present study aim to investigate the effects of dietary Gelsemium elegans alkaloids supplementation in Megalobrama amblycephala. A basal diet supplemented with 0, 5, 10, 20 and 40 mg/kg G. elegans alkaloids were fed to M. amblycephala for 12 weeks. The study indicated that dietary 20 mg/kg and 40 mg/kg G. elegans alkaloids supplementation could significantly improve final body weight (FBW), weight gain rate (WGR), specific growth rate (SGR), feed conversion ratio (FCR) and protein efficiency ratio (PER) (P < 0.05). The 20 mg/kg and 40 mg/kg G. elegans alkaloids groups showed significantly higher whole body and muscle crude protein and crude lipid contents compared to the control group (P < 0.05). The amino acid contents in muscle were also significantly increased in 20 mg/kg and 40 mg/kg groups (P < 0.05). Dietary 40 mg/kg G. elegans alkaloids had a significant effect on the contents of LDH, AST, ALT, ALP, TG, TC, LDL-C, HDL-C, ALB and TP in M. amblycephala (P < 0.05). Fish fed 20 mg/kg and 40 mg/kg dietary G. elegans alkaloids showed significant increase in complement 3, complement 4 and immunoglobulin M contents. The liver antioxidant enzymes (SOD, CAT and T-AOC) and MDA content significantly increased at 20 mg/kg and 40 mg/kg G. elegans alkaloids supplement (P < 0.05). The mRNA levels of immune-related genes IL-1ß, IL8, TNF-α and IFN-α were significantly up-regulated, whereas TGF-ß and IL10 genes were significantly down-regulated in the liver, spleen and head kidney of fish fed dietary supplementation with 20 mg/kg and 40 mg/kg G. elegans alkaloids. After challenge with Aeromonas hydrophila, significant higher survival rate was observed at 20 mg/kg and 40 mg/kg G. elegans alkaloids supplement (P < 0.05). Therefore, these results indicated that M. amblycephala fed a diet supplemented with 20 mg/kg and 40 mg/kg G. elegans alkaloids could significantly promote its growth performance, lipids and amino acids deposition, immune ability and resistance to Aeromonas hydrophila.

Induction of oxidative stress, apoptosis and DNA damage by koumine in Tetrahymena thermophila.

Koumine is a component of the Chinese medicinal herb Gelsemium elegans and is toxic to vertebrates. We used the ciliate Tetrahymena thermophila as a model to evaluate the toxic effects of this indole alkaloid in eukaryotic microorganisms. Koumine inhibited T. thermophila growth and viability in a dose-dependent manner. Moreover, this drug produced oxidative stress in T. thermophila cells and expressions of antioxidant enzymes were significantly elevated at high koumine levels (p < 0.05). Koumine also caused significant levels of apoptosis (p < 0.05) and induced DNA damage in a dose-dependent manner. Mitophagic vacuoles were present in cells indicating induction of autophagy by this drug. Expression of ATG7, MTT2/4, CYP1 and HSP70 as well as the MAP kinase pathway gene MPK1 and MPK3 were significantly altered after exposed to koumine. This study represents a preliminary toxicological evaluation of koumine in the single celled eukaryote T. thermophila.

Effects of fasting and re-feeding on mstn and mstnb genes expressions in Cranoglanis bouderius.

The myostatin (mstn) and myostatinb (mstnb) gene of Cranoglanis bouderius were cloned and sequenced and their expressions under nutritional restriction were characterized. The full cDNA sequences of mstn and mstnb were 1878 bp and 1928 bp, containing an open reading frame of 1170 bp and 1119 bp, which encoded 390 and 373 amino acids, respectively. The deduced mstn and mstnb sequence structures were similar to other members of TGF-ß superfamily, including the TGF beta pro-peptide, TGF beta domain, proteolytic processing site and nine conserved cysteines in the C-terminal. In addition, four mstn gene duplications were found in Cranoglanis bouderius. Sequence alignment and phylogenetic tree analyses indicated that the mstn gene and mstnb gene had a close relationship with Siluriformes fish, and the mstn and mstnb genes were roughly classified into two groups. RT-PCR analysis revealed that the mstn and mstnb were expressed in a variety of tissues in Cranoglanis bouderius although the mstn was highly expressed in skeletal muscle and the mstnb was mainly expressed in brain. We speculate that the mstn gene but not mstnb is likely to play a key role in managing muscle growth. A fasting-re-feeding experiment was used to evaluate the effects of starvation on mstn and mstnb expressions in juvenile Cranoglanis bouderius for 5 weeks. The result showed that the mstn and mstnb transcript levels varied among tissues. The mRNA expression levels of mstn in muscle, brain and liver gradually decreased during starvation and returned to the normal level after re-feeding. The mstnb mRNA levels in muscle, brain, liver, spleen, intestine and kidney increased during an early fast time but ultimately decreased with prolonged fasting time. The mstnb transcript levels in muscle, brain and liver increased significantly after re-feeding. In summary, the results supported that the mstn and mstnb may not be limited to control of muscle growth in fish but could also be involved in other biological functions.