| Golden pompano(Trachinotus ovatus),belongs to the Trachinotus and Carangidae,and is a euryhaline fish.Because of fast growth,high nutritional value,simple diet,and easy rearing,T.ovatus has become one of the most important cultured marine species in the Southeast coastal areas of China.Currently,the rearing methods of T.ovatus mostly adopt high stocking density cage culture.High stocking density will make fish under stress for a long time,which will affect the growth and welfare of fish and increase the risk of aquaculture management.In this paper,the effects of different stocking densities on the growth,physiology and biochemistry of juvenile T.ovatus are studied,hoping to provide some reference for the healthy breeding of golden pompano and improve the welfare of fish.In this study,juvenile T.ovatus was used as the research object.the experimental fish(9.75 ± 0.11 g)were randomly divided into three density groups.There were three replicates for each stocking density.The cage(1m × 1m × 1m)was stocked with 100,200,and 300 fish,respectively,which were labeled as low stocking density group(LD),medium stocking density group(MD),and high stocking density group(HD).The experiment lasted 10 weeks.The experimental results are as follows:(1)The experimental results showed that the growth performance decreased significantly and the feed conversion rate increased significantly of T.ovatus in the MD and HD(P < 0.05).No signifcant difference in specifc growth rate was found among the different groups(P > 0.05).The condition factor decreased with the increase of stocking density,and was the highest in the LD(P < 0.05).There was no significant difference in serum cortisol levels among the different groups(P > 0.05);the serum thyroxine levels in the LD and MD were significantly higher than those in the HD(P < 0.05).Serum glucose concentration increased significantly in the MD and HD(P < 0.05);total protein,total triglyceride,and total cholesterol concentrations decreased significantly in the HD(P < 0.05).Muscle component test results showed that the crude fat content was the highest in the LD(P < 0.05),while the moisture,ash and crude protein content had no significant difference among the different groups(P > 0.05).In this experiment,superoxide dismutase,glutathione peroxidase,and total antioxidant capacity increased with increasing stocking density.Serum lysozyme,C3 and C4 levels increased significantly in the HD(P < 0.05).The results of q RT-PCR showed that the expression level of HSP70 in liver,kidney,and brain was significantly increased with the increase of stocking density(P < 0.05),but the expression in spleen was not significantly different(P > 0.05).In general,higher stocking density has a negative impact on the growth,feed conversion rate and physiology and biochemistry of juvenile golden pompano;this means that farming costs will increase,which is not conducive to economic benefits and also reduces the welfare level of T.ovatus.(2)As a stress protein and important molecular chaperone,heat shock protein 90 has recently received considerable attention in aquaculture research,and it plays an important biological role in biological tolerance to various stressors.In this paper,the c DNA sequences of To GRP94,To HSP90α,and To HSP90β from T.ovatus were obtained for the first time.To GRP94 c DNA(Gen Bank accession No: MT120311)has a sequence length of 2711 base pairs(bp),and its open reading frame(ORF)contains 2394 bp and encodes 797 amino acids.To GRP94 amino acid sequence has a signal peptide sequence at the N-terminus,a heat shock hsp90 proteins family signature(Yk NKEIFLRE),five conserved signal regions of the HSP90 family,and an endoplasmic reticulum targeting sequence(KDEL).To HSP90α c DNA(Gen Bank accession No: MT120312)has a sequence length of 2429 bp,and its ORF contains 2190 bp,which can encode 730 amino acids.To HSP90α amino acid sequence has a heat shock hsp90 proteins family signature(Ys NKEIFLRE)and five HSP90 family conserved signal regions.To HSP90β(Gen Bank accession No: MN641479)has a sequence length of 2648 bp,of which the ORF contains 2184 bp,which is speculated to encode a protein containing 728 amino acids.To HSP90β amino acid sequence has one heat shock hsp90 proteins family signature(Ys NKEIFLRE)and five HSP90 family conserved signal regions.Sequence alignment and phylogenetic analysis showed that the amino acid sequences of these three genes are highly identical to the corresponding amino acid sequences known from other fish,and belong to the GRP94,HSP90α,and HSP90β subfamilies in the HSP90 family,respectively.By q RT-PCR detection,three genes were expressed in nine tissues tested.To GRP94 had the highest expression in gills tissue,followed by fins,kidney,and liver tissues,and the expression of other tissues is relatively low.To HSP90α had the highest expression in blood,followed by brain,kidney,and gills tissues.To HSP90β had the highest relative expression level in the intestine,followed by kidney,gills,fins,and liver tissues.Three genes are expressed in the all tissues being tested,suggesting that the three genes have extensive biological functions.All three genes were up-regulated to varying degrees in the corresponding tissues under stocking density chronic stress.The expression of To GRP94 in brain and muscle was significantly up-regulated in the HD group(P < 0.05),while its expression in the kidney was not significantly different between the groups(P > 0.05).To HSP90α expression levels in liver,kidney,and muscle were up-regulated with increasing stocking density,and the expression levels in the HD group were significantly higher than those in the LD and MD groups(P < 0.05).There was no significant difference in the expression levels of To HSP90α in the brain between the groups(P > 0.05).Similarly,the relative expression levels of To HSP90β in the kidney and brain increased significantly with increasing stocking density(P < 0.05),with the highest expression in the HD group.The relative expression of To HSP90β m RNA in muscle was significantly highest in the HD group(P < 0.05),while it was lowest in the MD group.These three genes are closely related to density stress,and high stocking density may cause chronic crowding stress on experimental fish. |
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