Effects Of Dietary Magnesium Level On The Growth Performance,Functional Organ Health And Flesh Quality,as Well As The Possible Mechanisms In Young Grass Carp (Ctenopharyngodon Idella)

This study investigated the effects of magneisum level on the growth performance,disease resistance,functional organ health and flesh quality,as well as the possible mechanisms in young grass carp（Ctenopharyngodon idella）,and determined the optimal dietary magnesium requirements for young grass carp.A total of 540 healthy grass carp（223.85±0.49 g）were fed six diets containing graded levels of magnesium(73.54,281.37,487.49,691.55,861.67 and1054.53 mg kg^-1 diet)for 60 days to investigate the effects of magnesium on growth performance and flesh quality of young grass carp.After growth trial,fish were challenged with Aeromonas hydrophila for 14 days and Flavobacterium columnare for 3 days to investigate the effects of magnesium on the immune function,apoptosis related parameters,antioxidant ability and tight junction protein as well as its potential mechanisms in intestine,head kidney,spleen,skin and gill of young grass carp.The results showed that compared with magnesium deficiency(73.54 mg magnesium kg^-1 diet),optimal magnesium level increased percent weight gain（PWG）,feed intake（FI）,feed efficiency（FE）and specific growth rate（SGR）（P<0.05）,suggesting that magnesium could improve the growth performance of young grass carp.Optimal magnesium level decreased enteritis,skin hemorrhage and lesion,and gill rot morbidity（P<0.05）,suggesting that magnesium could increase the disease resistance of young grass carp.The ability against enteritis,skin hemorrhage and lesion,and gill rot morbidity are related to the functional organs health of young grass carp.In this study,optimal magnesium level increased lysozyme（LZ）and acid phosphatase activities（ACP）,complement 3,complement 4 and immunoglobulin M（Ig M）contents in the intestine,gill,skin,head kidney and spleen of young grass carp（P<0.05）,up-regulated the m RNA levels of hepcidin,mucin2,liver-expressed antimicrobial peptide 2（LEAP-2）,β-defensin-1,anti-inflammatory cytokines,TGF-β1（except in the mid intestine and distal intestine）,TGF-β2（except in the spleen,mid intestine and distal intestine）,interleukin（IL）-4/13A,IL-4/13B,IL-10（except in the gill and intestine）,IL-11,inhibitor ofκBα（IκBα）,target of rapamycin（TOR）and ribosomal protein S6 kinases 1（S6K1）in the intestine,gill,skin,head kidney and spleen of young grass carp（P<0.05）,whereas down-regulated pro-inflammatory cytokines,TNF-α（except in the spleen）,IFN-γ2（except in the gill）,IL-1β（except in the head kidney）,IL-6,IL-8,IL-12 p35（except in the gill and intestine）,IL-12 p40（except in the head kidney）,IL-15（except in the skin）,IL-17 and nuclear factor kappa B（NF-κB）p65,c-Rel,IκB kinaseβ（IKKβ）,IκB kinaseγ（IKKγ）and e IF4E-binding proteins 1（4E-BP1）（except in the skin）,4E-BP2 m RNA levels in intestine,head kidney,spleen,skin and gill of young grass carp（P<0.05）,suggesting that magnesium could alleviate the inflammation and enhance immune function of grass carp partly through the NF-κB p65 and TOR signaling pathways;Meanwhile,optimal magnesium level decreased contents of malondialdehyde（MDA）,protein carbonyl（PC）,reactive oxygen species（ROS）and GSH,increased the capacities of anti-superoxide anion（ASA）and anti-hydroxy radical（AHR）and increased the activities and m RNA levels of antioxidant enzymes catalase（CAT）,manganese superoxide dismutase（Mn SOD）,glutathione peroxidase（GPx）,glutathione-S-transferase（GST）and glutathione reductase（GR）（P<0.05）,and up-regulated the m RNA levels of NF-E2-related factor 2（Nrf2）（P<0.05）,down-regulated the m RNA levels of Kelch-like-ECH-associated protein 1b（Keap1b）in intestine,head kidney,spleen,skin and gill of young grass carp（P<0.05）,suggesting that magnesium could enhance the antioxidant ability of grass carp;Meanwhile,dietary magnesium up-regulated the m RNA levels of B-cell lymphoma protein-2（Bcl-2）,inhibitor of apoptosis proteins（IAP）and myeloid cell leukemia-1b（Mcl-1b）（P<0.05）,whereas down-regulated the m RNA levels of cysteinyl aspartic acid-protease（caspase）-2,-3（except in the spleen）,-7,-8,-9（except in the spleen）,Fas ligand（Fas L）,apoptotic protease activating factor-1（Apaf-1）,Bcl-2 associated X protein（Bax）,p38 mitogen activated protein（p38 MAPK）and c-Jun N-terminal protein kinase（JNK）in intestine,head kidney,spleen,skin and gill of young grass carp（P<0.05）,suggesting that magnesium could inhibit the apoptosis,maintiaining structural integrity in intestine,head kidney,spleen,skin and gill of grass carp;Meanwhile,dietary magnesium down-regulated the m RNA levels of myosin light chain kinase（MLCK）and up-regulate the m RNA levels of tight junction protein occludin,zonula occludens（ZO）-1,occludin（except in the spleen）,claudin-b（except in the gill）,-c,-3c（except in the spleen and gill）,-f,-7（except in the intestine）,-11 and-12（P<0.05）,suggesting that magnesium could maintain the intercellular structural integrity in intestine,head kidney,spleen,skin and gill of grass carp.In addition,optimal magnesium level increased the contents of muscle crude protein,crude fat,hydroxyproline（Hyp）,composition of free amino acid and fatty acid as well as p H value（P<0.05）,decreased muscle water,cooking loss,shear force,lactate content,cathepsins B and L activities as well as ROS,PC and MDA contents（P<0.05）,whereas increased the activities and m RNA levels of manganese superoxide dismutase（Mn SOD）,catalase（CAT）,glutathione peroxidases（GPx）,glutathione-S-transferases（GST）and glutathione reductase（GR）as well as glutathione（GSH）content（P<0.05）,up-regulated the m RNA levels of Nrf2,TOR,S6K1,casein kinase 2α（CK2a）and CK2β,and down-regulated the m RNA level of Keap1a and 4E-BP（P<0.05）,which enhanced the flesh quality of fish.As stated above,optimal magnesium level could increase the growth performance and ability of against enteritis,skin hemorrhage and lesion and gill rot morbidity.The increased ability against enteritis,skin hemorrhage and lesion and gill rot morbidity of young grass carp are related to the enhanced functional organs health with the supplemention of dietary magnesium.Optimal magnesium level enhanced the intestinal immune,body immune and gill immune function of young grass carp were related to the increased antibacterial compounds production and attenuated inflammation under the regulation of down-regulated pro-inflammatory cytokines which is partially related to NF-κB signalling and up-regulated anti-inflammatory cytokines which is partially related to TOR signaling in intestine,head kidney,spleen,skin and gill of young grass carp.Dietary magnesium enhance the structural integrity in intestine,head kidney,spleen,skin and gill of young grass carp which is partially related to,（1）inhibit apoptosis which is partially related to up-regulated anti-apoptosis factors and down-regulated pro-apoptosis factors gene expression,（2）enhance the antioxidant ability which is partially related to the up-regulation of gene expression and activities of antioxidant enzyme under the regulation of Keap1/Nrf2 signalling,（3）enhance the intercellular structural integrity which his partially related to the gene expression of tight junction protein under the regulation of MLCK signaling.Meanwhile,magnesium improve flesh quality which was partialy related to the increased physical quality,nutrition quality and antioxidant ability in grass carp.In this study,based on the growth performance,ability against enteritis,skin hemorrhage and lesion and gill rot morbidity,the optimal magnesium level for young grass carp（223-757g）were calculated to be 770.38,836.68,813.12 and 836.11 mg kg^-1.