| China has vast saline-alkaline land resources with an area of 9.91×107 hm2,which is widely distributed in the northeast,north,northwest and inland areas of China,including 19 provinces,municipalities and autonomous regions,most of these land are in a barren state.Saline-alkaline water has the characteristics of high p H,high carbonate alkalinity,high ionic coefficient and imbalance of main ion proportions,which make it difficult for most aquatic organisms to adapt,and aquaculture production is unstable.The rational development of saline-alkaline water resources and the use of saline-alkaline water for aquaculture will help solve the shortage of freshwater resources in fisheries and increase aquaculture production.Largemouth bass(Micropterus salmoides),belongs to the order Perciformes,Porcoidei,Cehtrachidae,and Micropterus.It is a high-quality freshwater fish with the advantages of strong adaptability,fast growth,easy to catch,and short farming cycle.In addition,the meat is delicious and tender,without intermuscular spines,which is welcomed by the market and consumers.Since its introduction in the 1980 s,it has become an important economic fish in Chinese freshwater aquaculture industry.Largemouth bass has been successfully cultured in some saline-alkaline waters such as Ningxia and Tianjin,but its adaptation range of and mechanism to saline-alkaline are still unclear.In this study,fish growth performance,muscle quality,physiological indicators,and gene expression will be studied in the saline-alkaline water environment,in order to provide a theoretical basis for farming Largemouth bass in saline-alkaline aquaculture.In this study,the growth performance of largemouth bass in a high saline-alkaline environment was studied,and the half-lethal concentration of carbonate and alkalinity of largemouth bass at 48 h was determined.The salt water group(SW,salinity 7.50±0.07,carbonate alkalinity 1.81±0.12 mmol/L),alkaline water group(AW,salinity 0.35±0.02,carbonate alkalinity9.96±0.03 mmol/L),and fresh water control group(FW,salinity 0.13±0.01,carbonate alkalinity1.82±0.11 mmol/L)were set to comparatively study the growth parameters,physiology parameters and muscle texture characteristic index of largemouth bass under long-term saline-alkaline stress.The results showed that the 48-hour carbonate alkalinity semi-lethal concentration of largemouth bass was 29.92±3.90 mmol/L,and it could survive safely in a water environment with a salinity below 10.After 105 days of farming experiment,there are no significant differences of survival rate and final weight among different groups;the specific growth rate(SGR)showed a regular variation.During 15d-45 d and 60d-75 d,SGR continued to decrease,while increased during 45d-60 d and 75d-90d;The condition factors of largemouth bass were less than 3 in all groups with FW>AW>SW.In AW group,the blood ammonia of largemouth bass within 24 hours showed an increase firstly,then a decreased,and finally stabilized.In SW group,the osmolality of largemouth bass was 319.53±29.51 m Osm/kg,comparing a lower level as 300.00±16.44 m Osm/kg in FW group.Largemouth bass raised in saline-alkaline water had better texture characteristics.Largemouth bass raised in SW had a higher muscle hardness with 34.70±4.86 N,while it had a higher springiness with 1.06±0.10 mm in AW,the plasma Na+ concentration of largemouth bass in the saline group was the highest(158.87±2.03 mmol/L);the saline group had the highest plasma carbon dioxide partial pressure(15.23±2.55 mm Hg)and lowest blood p H(7.16±0.07);there was no significant difference in blood gas index between alkaline water group and fresh water groupThe effects of alkalinity stress on the transcriptome of the brain,stomach,and head kidney of largemouth bass were studied,setting 6 experimental groups freshwater brain(FDB),freshwater stomach(FDS),freshwater head kidney(FDHK),alkaline water brain(ADB),and alkaline water stomach(ADS),alkaline water head kidney(ADHK).Largemouth bass transcriptome data statistics showed that FDB,FDS,FDHK and ADB,ADS,ADHK groups obtained 45205260,47042702,47042702 and 46779792,47360200 and 45604936 raw reads respectively.After filtering the original sequences,41749736(FDB),40805442(FDS)and43858828(FDHK),43674482(ADB),44345160(ADS),43414896(ADHK)clean reads were obtained.Q30 were 93.82%(FDB),94.44%(FDS),93.95%(FDHK),93.65%(ADB),94.42%(ADS),and 94.11%(ADHK),respectively,all of which were greater than 90%,indicating that the data were accurate.FDB vs.ADB obtained a total of 1962 differentially expressed genes(DEGs),including 1289 down-regulated genes and 673 up-regulated genes;FDS vs.ADS obtained a total of 781 differentially expressed genes,121 up-regulated genes,and 630 significantly down-regulated genes;FDHK vs.ADHK obtained a total of 1264 differentially expressed genes,696 up-regulated genes and 568 down-regulated.The differential genes obtained by alkalinity stress were enriched and classified by the KEGG database,and it was found that alkalinity stress could affect the expression of genes related to environmental information processing,organic systems and metabolism.Among them,alkalinity stress led to the significant up-regulation of brain genes related to active ligand-receptor interaction,calcium signaling pathway,ECM-receptor interaction,MAPK signaling pathway,and Wnt signaling pathway in brain tissue,gastric glyceride metabolism,cysteine and methionine metabolism,glycine,serine and threonine metabolism,neomycin,kanamycin and gentamicin biosynthesis,starch and sucrose metabolism related genes were significantly up-regulated;alkalinity stress could affect the expression of head kidney,including MAPK signaling pathway,cytokine-cytokine Receptor interaction and PPAR signaling pathway related genes were significantly up-regulated.The genes related to environmental information processing such as slc8a4 a,camk2a,LOC119884188,chrna6,il15 ra,il10ra,LOC119899106,LOC119882624,etc.,and amino acid metabolism-related genes such as LOC119892499,LOC119918947,etc.,LOC119908822,hspb1 and other immune-related genes can be used as candidate genes for largemouth bass to respond to alkalinity stress.The expression of genes related to osmotic regulation,ammonia nitrogen excretion and acid-base regulation in gills,kidneys and intestinal tissues of largemouth bass under long-term saline-alkaline stress were studied.Largemouth bass has higher expression of NKA in gill tissue in saline environment,mainly performing osmotic regulation and ion regulation.Kidney has higher expression of Rh family proteins in alkaline water environment,mainly excreting ammonia.The expression of various proteins in the intestine increased in the alkaline water environment,and participated in the process of ion transport and nutrient transport.After a 105-day saline-alkaline aquaculture experiment,the largemouth bass showed good growth performance in saline-alkali water.The largemouth bass had strong physiological regulation,gill tissue osmotic regulation and ion regulation,and the kidneys mainly performed ammonia excretion.Function,the intestine is involved in the process of ion transport and nutrient transport,and the expression of signal transduction genes in brain tissue increases;the expression of digestive and immune genes in gastric tissue increases,and the expression of immune-related genes in head and kidney tissue increases.This paper has 14 figures,10 tables,and 122 references. |
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