Effects And Underlying Mechanism Of Dietary Mannan Oligosaccharides On Immune And Disease Resistance Of Haliotis Discus Hannai Ino

Haliotis discus hannai Ino is one of the most important economic shellfish in China.Bacterial and viral infection often lead to high mortality and serious economic loss of cultured abalone.Among them,Vibrio parahaemolyticus is the main pathogen causing abalone disease.Mannan oligosaccharides（MOS）can promote the growth of beneficial microbes in the gut.MOS is a commonly used feed supplement for fish and crustaceans.Many studies have shown that MOS can significantly improve the growth,immunity and disease resistance of aquatic animals,However,not all aquatic animals have a positive response to MOS.In order to explore the specific action mechanism of MOS against Vibrio of Haliotis discus hannai Ino,the main contents and results are as follows:1.Four groups of multifunctional abalone diets were prepared by adding 0.0g/kg,0.4g/kg,0.8g/kg and 1.6g/kg Actigen ^TM（MOS 12%）,and were named A0,A4,A8 and A16,respectively.A 100 day culture experiment of Haliotis discus hannai Ino was carried out in an indoor circulating water system.The results showed that specific growth rate（SGR）and shell increase rate（SIR）of abalone in A8 group were significantly higher than those in other groups（p<0.05）,there was no significant difference between groups A4,A16 and control group（p<0.05）.Dietary MOS significantly increased the activities of AKP and ACP in hepatopancreas and serum of juvenile abalone（p<0.05）,but had no significant effect on hemocytosis and respiratory burst（p<0.05）.The lysozyme activity of abalone in A4 group was significantly higher than that in other groups（p<0.05）,but there was no significant difference between A8and A16 groups（p<0.05）.Transcriptome analysis showed that there were 2507,600and 683 differentially expressed genes（DEGs）in A4,A8 and A16 treatment groups compared with the control group A0.Compared with the control grow,up-regulated DEGs were mainly enriched in cytoplasmic matrix and chitin metabolism,and involved in ribosome,glutathione metabolism and phagosome KEGG pathways.Down-regulated DEGs are mainly enriched in GO items such as tetrapyrrole binding,heme binding and other ion binding,and participate in KEGG pathways such as carbohydrate and fatty acid metabolism.In addition,compared with A0,perlucin and collagen VI in A4,A8 and A16 were significantly up-regulated,suggesting that MOS can promote the growth and development of abalone by improving shell growth,stimulating collagen synthesis and promoting connective tissue growth.2.After fasting for 48 hours,the abalone in the above culture test was injected intramuscularly（100μL bacterial suspension with the final concentration of 4×10⁷cfu/m L）and immersion stress（final concentration is 4×10⁷CFU/m L,3 ml bacterial suspension in each aquarium）was used to challenge abalone for 56 hours.Count the number of abalone deaths every 8h and calculate the cumulative mortality.The gill tissues of each group（A0,A4,A8 and A16）before and after 8h/48h challenge were analyzed by transcriptome sequencing.The results showed that during 56h of Vibrio parahaemolyticus challenge,the cumulative mortality of abalone treated with MOS was significantly lower than that of control A0 group at all time points.At 56h of challenge,A8 group had the lowest cumulative mortality（about 10%）,which was significantly lower than A0（49%）,A4（30%）and A16（28%）（p<0.05）.Without MOS addition,compared with before challenge,there were 8322 DEGs in gill tissue of abalone after challenge（8h and 48h）.The down-regulated DEGs were enriched in some metabolic pathways,ABC transporters and cell adhesion related pathways.However,DEGs in NOD-like receptor signaling pathway and apoptosis were significantly up-regulated.Analysis of sequencing results of A0,A4,A8 and A16treatment groups at 0h,8h and 48h challenge showed that:compared with A0 group,A4,A8 and A16 treatment groups had 301,292 and 1184 DEGs at 8h challenge,respectively.Among them,571 up-regulated genes were mainly enriched in GO items such as peptidyl proline isomerism,peptidyl proline cis-trans isomerase activity and KEGG pathways such as metabolic pathways.1207 down-regulated genes were mainly enriched in GO items such as sugar and lipid metabolism and glutathione metabolic pathway.Under 48h vibrio challenge,compared with A0 group,there were 2414,304and 4741 DEGs in A4,A8 and A16 treatment groups,respectively.Among them,5217up-regulated genes were mainly enriched in purine nucleotide synthesis,small molecule synthesis and other GO items,as well as KEGG pathways such as cell adhesion,ABC transport,RNA transport and apoptosis.2242 down-regulated genes are mainly enriched in GO items such as peptide transport and KEGG pathways such as metabolism and oxidative phosphorylation.After vibrio challenge,MOS could enhance the expression of pattern recognition receptors to actively respond to vibrio invasion and carry out immune defense.Promote the MAPK pathway,induce cell proliferation and differentiation for self-repair;Meanwhile,GST and HSP90 were up-regulated to maintain redox equilibrium.In conclusion,dietary MOS supplementation can significantly improve the growth and immune performance of abalone,and improve the survival rate of abalone when challenged with V.parahaemolyticus,and the optimal supplemental level is 0.8g/kg diet.