Effects Of Dietary Phosphorus On Growth Performance,the Health Status Of Intestinal,Gill And Body,and Flesh Quality Of Young Grass Carp(Ctenopharyngodon Idella) And The Mechanisms

This study was designed to determine the effects of phosphorus on the growth prefermance,disease resistance,intestinal health,gill health,body health and flesh quality,as well as its mechanisms in young grass carp(Ctenopharyngodon idella)by growth trial and challenge trial.A total of 540 young grass carp(256.22 ± 0.60 g)were randomly distributed into six groups of each three replicates,and fed six diets containing graded levels of available phosphorus at 0.95,2.46,3.96,5.68,7.10 and 8.75 g/kg diet(totle phosphorus levels were 2.3,4.0,5.6,7.6,9.2 and 11.0 g/kg)for 60 days.Subsequently,two challenge tests were conducted by injection of Aeromonas hydrophila for 14 days and Flavobacterium columnare for 3 days to investigate the influences of phosphorus on disease resistance,intestinal health,gill health and body health and its meachimis by exploring the effects of phosphorus on antibacterial substances,inflammation,antioxidant,apoptosis and tight junction protein in the intestine,gill,skin,head kidney and spleen of young grass carp.The results of growth trial showed that,compared with dietary phosphorus deficiency(0.95 g/kg diet),optimal level of phosphorus increased percent weight gain(PWG),feed intake(FI),feed efficiency(FE)and specific growth rate(SGR)(P<0.05),indicating that phosphorus improved the growth performance of young grass carp.Phosphorus increased the contents of protein,essential amino acids,flavor amino acids,PUFA and hydroxyproline and pH value,as well as decreased shear force,cooking loss,lactate content,cathepsin B and L activities and saturated fatty acid content in the muscle of young grass carp,suggesting that optimal phosphorus level could increase fish flesh quality.Dietary phosphorus increased Kelch-like ECH-associating protein 1/Nrf2 signaling to promote gene expressions and activities of antioxidant enzymes such as copper/zinc superoxide dismutase(CuZnSOD),and decreased oxidative injury.Meanwhile,optimal level of phosphorus decerased enteritis,gill rot and skin hemorrhage and lesions morbidity(P<0.05),suggesting that phosphorus could enhance the ability to combat enteritis,gill rot and skin hemorrhage and lesions of young grass carp which are related to the health status of intestine,gill and body,respectively.The results of this study showed that optimal phosphorus level promoted the lysozyme(LZ)and acid phosphatase(ACP)activities and the complement 3(C3),C4 and immunoglobulin M(IgM)contents as well as the mRNA levels of antimicrobial peptides[liver expressed antimicrobial peptide(LEAP)-2A,-2B and ?-defensin-1]in the intestine,gill,skin,head kidney and spleen of young grass carp,suggesting that phosphorus could enhance antimicrobial ability of fish.Optimal phosphorus level down-regulated the gene expressions of I?B kinase ?(IKK?),IKK? and nuclear factor kappa B(NF-?B)p65 and up-regulated inhibitor of ?B?(I?B?)leading to the down-regulation of pro-inflammatory cytokines interleukin 1?(IL-1?)(except in the distal intestine),IL-6(except in gill),IL-15,IL-17D(except in gill)and interferon ?2(IFN-?2)gene expressions.In addition,optimal phosphorus level up-regulated the gene expressions of TOR and S6K1 and down-regulated 4E-BP causing the up-regulation of anti-inflammatory cytokines IL-4/13A,IL-10,IL-11,TGF-?1 and TGF-?2 gene expressions,indicating that phosphorus inhibited the inflammation in the mucosal immune and body immune organs of fish,which might be partially related to NF-?B and TOR signling.A futher study revealed that optimal level ofphosphorus inhibited Fas ligand/caspase-8 death receptor pathway and Bcl-2 associated X protein(Bax)family/apoptotic protease activating factor-1(Apaf-1)/caspase-9 mitochondria pathway,and down-regulated caspase-3,-7 gene expression resulting the inhibition of cell apoptosis and the enhancement of cellular integrity;phosphorus promoted Nrf2 signaling pathway to increase the gene expressions and activites of antioxidant enzymes manganese superoxide dismutase(MnSOD),catalase(CAT)(but not in spleen),glutathione peroxidase(GPx)1a,1b and glutathione S-transferases(GST)R,O1,O2,P1,P2,and then decreased reactive oxygen species(ROS),malondialdehyde(MDA)and protein carbonyl(PC)contents.This suggested that phosphorus increased the ability of antioxidant,and decreased oxidative damage in the intestine,gill,skin,head kidney and spleen of fish.Moreover,the down-regulated of signling molecule myosin light-chain kinase(MLCK)mRNA levels by dietary phosphorus increased the mRNA levels of zonula occludens,occludin(except in the proximal intestine and mid intestine),claudin-c,-f,-7 and-11 and decreased claudin-12,indicating that phosphorus could maintain the intercellular structural integrity in the intestine,gill,skin,head kidney and spleen of fish.In conclusion,optimal phosphorus could improve growth performance and increase antioxidant ability to improve muscle physicochemical characteristic,flavor amino acids,healthcare function fatty acids,and thus enhance fleah quality of young grass carp.Those are related to the enhancement of the ability to combat enteritis,gill rot and skin hemorrhage and lesions,which may be associated with intestinal health,gill health and body health,as is displayed in following aspects:(1)Optimal phosphorus level increased antibacterial ability by promoting the activities and contents of major antibacterial compounds,and up-regulating the mRNA levels of antibacterial peptides;modulated NF-?B and TOR signaling to down-regulate the mRNA levels of pro-inflammatory cytokines and up-regulate anti-inflammatory cytokines,leading to the attenuation of inflammation in the intestine,gill,skin,head kidney and spleen of young grass carp.(2)Optimal phosphorus level improved the mRNA levels and activities of antioxidant enzymes(like SOD)to enhance the ability of scavenge free radical and decrease oxidative damage partly by Nrf2 signaling,and decreased the death receptor pathway and mitochondria pathway to inhibite apoptosis,leading to the normal cellular integrity;and enchance the barrier function of tight junction protein to maintain the intercellular integrity in the intestine,gill,skin,head kidney and spleen of young grass carp.Finally,based on PWG,the ability to combat enteritis,gill rot and skin hemorrhage and lesions,the dietary available phosphorus requirements for young grass carp(254-898 g)are estimated to be 4.10,4.68,5.77 and 4.13 g/kg diet,respectively.