| Yellow drum(Nibea albiflora)is an important marine fish for aquaculture.The southeast coast of China often suffers from the cold current in winter due to global climate change,resulting in freezing death of a large number of cultured yellow drum causing huge economic losses,and greatly restricting the development of breeding industry.Therefore,in this study,the response mechanisms in physiological level and transcriptome level of yellow drum under low temperature and starvation stresses were studied.There were four groups in the experiment which took 30 days: 16°C feeding group(16fed,control),16°C unfed group(16unfed),8°C unfed group(8unfed)and 8°C feeding group(8fed).The research results are as follows:1.By measuring the growth performance data of the yellow drum,we found that the body weight,total length and body length of the 8unfed group were significantly lower after the experiment compared with those before the experiment(P < 0.05);there was no significant decrease of 16 unfed group after the experiment in body weight,but the total length and body length were significantly increased compared with those before the experiment(P < 0.05).The liver/body ratio of the 16 fed group was significantly higher than that of 16 unfed and 8unfed(P < 0.05).The results showed that the growth and development of yellow drum were seriously affected by low temperature and starvation.By observing the liver histology of yellow drum,we found that the liver tissue cells of starvation stress group were irregular,closely arranged and disordered,some cells were diffuse,the whole liver cells were deeply stained,and the liver cells of cold stress group showed vacuolation,cell boundary disorder,nuclear atrophy and even dissolution,indicating that low temperature and starvation could damage the liver tissue of yellow drum.2.By measuring the relative gene expression levels of antioxidant genes in the liver of the yellow drum,we found that the relative expression of the antioxidant genes(copper-zinc superoxide dismutase(Cu,Zn-SOD),manganese superoxide dismutase(Mn-SOD),iron superoxide dismutase(Fe-SOD)and catalase(CAT))in the liver of the low temperature and starvation stress group was lower than that of the control group(16fed).The result consistent with the results of enzymatic activity of superoxide dismutase(SOD)and catalase(CAT).The glutathione peroxidase(GSH-Px)gene expression level showed different degrees of differential expression between the groups at most sampling points,and a higher expression level was observed in the 16 unfed group.In addition,the activities of immunoregulatory enzymes(alkaline phosphatase(AKP)and acid phosphatase(ACP))were also inhibited in the hypothermia and starvation stress groups.The results showed that low temperature and starvation stress significantly inhibited the activity and gene expression of some enzymes in the liver of yellow drum.It is speculated that low temperature and starvation stress can inhibit the immunity of yellow drum.3.The transcriptome sequencing of liver tissues from d22 of each group was carried out by RNA sequencing technology.A total of 62536 unigenes were spliced,with a total length of 92,038,838 bp and an average length of 1471 bp.Unigene’s database annotation results are as follows: 29924(47.85%)genes are annotated to the NR library,25257(40.39%)genes are annotated to the Swissprot library,18126(28.98%)genes are annotated to the KEGG library,and 19677(31.47%)genes are Annotated to the KOG library,27724(44.33%)genes were annotated into the eggNOG library,23077(36.90%)genes were annotated into the GO library,and 47(0.08%)genes were annotated into the Pfam library.The reads were compared to Unigene with a ratio of 95.36 to 96.61%.There are six differential groups,and the number of differential genes detected is 2259(16fed vs 8unfed),33(8unfed vs 8fed),1912(16fed vs 8fed),999(16fed vs 16unfed),432(16unfed vs8fed)and 326(16unfed vs 8unfed).Through KEGG and GO cluster analysis,it was found that differentially expressed genes under low temperature and starvation stress were mostly enriched in "carbobolism metabolism","energy metabolism","translation","signal molecule interaction","immune system" and " Transport and catabolism" and other signaling pathways.These signaling pathways can be functionally classified into: signal transduction,energy metabolism,and the immune system.In addition,eight differentially expressed genes were selected from the transcriptome for qRT-PCR validation,and the results confirmed that the RNA-seq results were reliable.This study preliminarily explored the response mechanism of the yellow drum at physiological and transcriptome level under low temperature and starvation stress.The results showed that low temperature and starvation stress could significantly inhibit the growth,damage its liver cells,and affect the activity and expression of antioxidant enzymes in its liver.In addition,through the study of transcriptome expression profiles,it was also found that low temperature and starvation stress may affect the metabolism and immune system of yellow drum.The results will be helpful to understand the physiological response mechanism of yellow drum during winter under low temperature and starvation,and provide theoretical basis for breeding cold-tolerant varieties of yellow drum. |
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