Immune Response Of Skin Mucosal Immune System Of Siniperca Chuatsi To Ambient Temperature And Aeromonas Hydrophila Inactivated Vaccine

Fish skin gland layers of the skin contains abundant glandular cell,can secrete mucus covered in body surface,constitute its contact with the outside world the first line of defense,the mucus contains a lot of antibacterial substances,such as antibacterial peptide,lysozyme,protease and lectin material can resist bacteria and parasites invade,skin mucous also in fish to maintain osmotic pressure,condensation,precipitation water suspended material,Moistening the skin and so on plays a key role.Therefore,the aquatic environment and biological environment on the surface of skin mucous membrane immune system response has an important influence,under this background,take Siniperca chuatsi as the research object,this paper carried out a series of physiological in-depth studies on the skin and mucosal immune system of Siniperca chuatsi after being exposed to different ambient temperatures and soaking inactivated vaccine of Aeromonas hydrophila by using histological,biochemical and molecular methods.Firstly,the types and number of skin mucus cells of Siniperca chuatsi under different ambient temperatures were observed,and the effects of ambient temperature on the activities of non-specific immune-related enzymes in skin mucus and the expression levels of immune-related genes in skin were measured.The changes of antibody level and activity of non-specific immune-related enzymes in the mucus of Siniperca chuatsi and the immune response of the expression level of immune-related genes in the skin of Siniperca chuatsi after immune-immersion of Aeromonas hydrophila inactivated vaccine provide basic data for the skin and mucosal immune system of Siniperca chuatsi to deal with the changes of external temperature and the invasion of pathogenic bacteria.The results are as follows:In order to understand the effects of different temperature conditions(10,20,30 ℃)on the mucus cell types and immune factors on the skin surface of Siniperca chuatsi,the distribution and types of mucus cells on the skin surface of Siniperca chuatsi were observed by AB-PAS staining.The staining results showed that there are four types of mucus cells on the skin surface of Siniperca chuatsi.The number of mucus cells in the head is the most,followed by the trunk and the tail.Compared with 10 ℃ group,typeⅡ increased the most in 20 ℃ and 30 ℃ groups,followed by type Ⅳ,type Ⅰ showed a slightly decreasing trend,and type Ⅲ showed irregular change.q PCR and double-antibody sandwich enzyme-linked immunoassay were used to detect the changes of gene expression levels of skin surface immune factors(Ig M,IL-1β,Hepcidin)and the changes of enzyme activities of SOD,ACP and LYS.The results showed that the expression levels of Ig M gene and Hepcidin gene were up-regulated with increasing temperature.Compared with the 10 ℃ group,the expression levels of IL-1β gene were significantly up-regulated in the 20 ℃ group and down-regulated in the 30 ℃ group.The SOD activity in 30 ℃ group was significantly higher than that in 10 ℃ and 20 ℃groups,while the ACP and LYS activities were slightly higher than those in 10 ℃ and20 ℃ groups.The above results indicate that the skin mucus cell types and number of siniperca chuatsi and immune-related factors are significantly affected by ambient temperature.In order to study the immune response of skin mucosal immune system of Siniperca chuatsi after immersion immunization with Aeromonas hydrophila inactivated vaccine,the Aeromonas hydrophila inactivated vaccine was prepared by formalin inactivation method.The healthy siniperca chuatsi was immunized by immersion method,and the antibody levels in the skin mucus of siniperca chuatsi and the changes of SOD,ACP and LYS enzyme activities in the mucus were measured after immunization.And expression levels of skin immune factors Ig M,IL-8,Hepcidin and MHC Ⅱ genes.The results showed that the specific antibody level in the skin mucus of Siniperca chuatsi reached the peak at 11 d,and was significantly higher than that of the control group within 4～14 d(P<0.05).The activity of LYS and SOD in skin mucus increased and reached the peak in 2～4 d after soaking,and then decreased with the extension of time.The overall change trend of ACP and ALP was relatively gentle,and reached the peak in 2-4 days,and there was no significant difference between the activity and the control group in other time periods(P>0.05).The peak time of immune-related genes in skin was relatively late,the expression levels of Ig M,IL-8 and Hepcidin reached the peak on 7 d,the expression levels of MHC Ⅱ reached the peak on11 d,and the expression levels in other time periods were higher than those in the control group.The results showed that the immersion of Aeromonas hydrophila inactivated vaccine significantly affected the antibody levels and non-specific related enzyme activities in the mucus of Siniperca chuatsi and the expression of immune-related genes in the skin,so as to enhance the immune protection ability of the fish.