Effects Of Curcumin On Antioxidant,Immune Function And Intestinal Flora Of Procambarus Clarkii

Procambarus clarkii is an important economic aquaculture species in China,and with the increase of culture production,area and density year by year,disease outbreaks,germplasm degradation and water environment pollution are increasing,hindering the healthy development of Procambarus clarkii culture industry.In recent years,herbal medicines and their extracts have been favored in disease prevention and control of aquaculture animals because of their excellent characteristics such as good efficacy,high safety,no pollution to the environment and low drug resistance.Curcumin has various biological functions such as antioxidant,anti-inflammatory and antiviral,to investigate the biological role of curcumin in the feeding of Procambarus clarkii.In this experiment,different levels of curcumin were added to the feed for indoor rearing of Procambarus clarkii,and the optimum curcumin addition was screened according to the effects of different curcumin additions on the hepatopancreas structure,antioxidant enzyme activity,immune function and intestinal flora of Procambarus clarkii.The optimum amount of curcumin was applied to the pond culture of Procambarus clarkii,and the effects of curcumin on the hepatopancreas structure,antioxidant enzyme activity,immune function and intestinal flora of pond culture of Procambarus clarkii were investigated to provide a theoretical basis for the application of curcumin in the healthy culture of Procambarus clarkii.The results of the study are as follows.1.Effects of different levels of curcumin in feed on antioxidant,immune function and intestinal flora in the indoor culture of Procambarus clarkiiFirstly,450 Procambarus clarkii with an initial weight of（12.74±0.39 g）were selected and randomly divided into 5 groups of 3 replicates each with 30 fish.The results of HE staining showed that the Cur4 and Cur6 groups had more uniform distribution among hepatic tubules,reduced the number of flattened hepatic tubule morphology,and increased the number of hepatopancreas luminal star-shaped cells.The hepatopancreatic tissue was better developed.Feeding curcumin significantly increased the enzyme activities of T-AOC,GSH-PX,SOD,CAT,LZM and PO（p<0.05）,with the Cur4 group having the highest T-AOC,SOD,GSH-PX and PO enzyme activities and the best effect.The expression of SOD,CAT,GSH-PX,and PO genes were found to be significantly up-regulated by fluorescence quantitative PCR（P<0.05）,and feeding curcumin enhanced the antioxidant capacity and immune function of Procambarus clarkii.The gut flora was analyzed by high-throughput 16s r RNA sequencing technology,and principal component analysis revealed that the composition of gut flora in the 7d Cur4 group was different from that of the control group,the composition of gut flora in the 21d Control and Cur2,Cur4,Cur6 and Cur8 groups was changed,and the composition of gut flora in the 28d Control and Cur2,Cur4,Cur6 and Cur8 groups was the same.Alpha diversity analysis revealed that the ACE index,Chao1 index,Shannon index,and Simpson index did not produce significant differences between Control and Cur2,Cur4,Cur6,and Cur8 groups at different time points（p>0.05）.Control and Cur2,Cur4,Cur6,and The distribution of intestinal flora at the phylum level in the Cur8 group mainly consisted of Firmicutes,Proteobacteria,Bacteroidetes,and Tenericutes;the distribution of intestinal flora at the genus level showed that the dominant genera in the Cur2,Cur8,and Control groups were the most abundant genus in Cur4 group was Shewanella（25.84%）,and the abundance of pathogenic bacteria in Cur2,Cur8 and Control groups was reduced;the most abundant genus in Cur6 group was Candidatus Bacilloplasma（38.97%）.The highest abundance in the Cur6 group was Candidatus Bacilloplasma（38.97%）,and the composition of the dominant genus at the genus level was changed.The results showed that different additions of curcumin were beneficial to the hepatopancreas structure,promoted the antioxidant capacity and immune function of Procambarus clarkii,improved the intestinal flora and reduced the abundance of pathogenic bacteria.The Cur4 group had better hepatopancreas tissue structure and the highest activity of antioxidant and immune-related enzymes,which were most effective in reducing the number of pathogenic bacteria.2.Effect of optimal curcumin addition on pond culture of Procambarus clarkiiBy comparing the experimental results of different additions of curcumin on the hepa topancreas tissue structure,antioxidant enzyme activity,immune function and intestinal fl ora of Procambarus clarkii,0.04%curcumin（Cur4）was selected for the experiment of p ond culture of Procambarus clarkii,the control group was the group without curcumin addition（Control）,two ponds were used as the experimental group and two ponds were use d as the control group for 3 weeks of feeding experiments.p H,DO,NO₂^-,NH₃-N,H₂S ph ysicochemical parameters of the cultured water were within the safe range for Procambar us clarkii culture and suitable for the growth of Procambarus clarkii.HE staining results showed that the lumen of the liver tubules of Procambarus clarkii in the 0.04%curcumin group had an obvious stellate structure,the liver tubules had a round structure,the numbe r of flattening was reduced,and the liver tubules The lumen of the liver tubules in the 0.04%curcumin group showed obvious stellate structure,rounded structure,reduced number of flattened tubules and better structural development compared with the control group.T-AOC,GSH-PX,SOD,CAT and PO activities were significantly higher in Procambarus c larkii after feeding 0.04%curcumin than in the control group（p<0.05）,while no signific ant difference occurred in LZM activity.The results of high-throughput 16s r RNA sequen cing showed that the structure of the intestinal flora in the Cur4 group was changed（p<0.05）,and the results of Alpha diversity analysis showed that the ACE index and Simpson i ndex in the Cur4 group were significantly higher（p<0.05）,which improved the richness and diversity of the intestinal flora.Control and Cur4 groups were mainly bacterium＿PM5＿3,ZOR0006,Anaerorhabdus,Weeksellaceae,Enterobacteriaceae.0.04%curcumin gro up Weeksellaceae,Enterobacteriaceae decreased the abundance of pathogenic bacteria.In summary,the following conclusions were drawn:（1）Different additions of curcumin improved the antioxidant and immune enzyme activity in the hepatopancreas and hemolymph of Procambarus clarkii,and improved the structure of the intestinal flora,with the best effect of 0.04%curcumin addition.（2）0.04%curcumin in pond culture of Procambarus clarkii can improve the antioxidant capacity and enhance the immune function,improve the intestinal flora,and reduce the abundance of pathogenic bacteria.