| Iridovirus disease of Larimichthys crocea is one of the most serious diseases that harm the breeding industry of large yellow croaker at present.It is mostly prevalent in the summer heat season,and usually occurs in young fish with a mortality of more than 50%.Currently,there is no effective targeted treatment for this disease.In recent years,in the actual production of large yellow croaker,starvation therapy is often used in the treatment of large yellow croaker iridovirus infection disease.This experiment by simulating clinical large yellow croaker iridovirus disease starvation therapy test,established a large yellow croaker artificially infected iridovirus group,respectively a week of continuous starvation and feeding,real-time fluorescence quantitative PCR was used to study the virus load and the expression of related immune factors in different tissues at the molecular level.Meanwhile,the indexes of immunoenzymes and antioxidant enzymes were determined and analyzed.The results were as follows:(1)According to the gene sequence recommended by the OIE diagnostic manual for aquatic animals,constructing the iridovirus SYBR Green?real-time fluorescent quantitative PCR detection method,its minimum detection limit was 10~2copies/ng DNA,and the correlation coefficient of plasmid standard curve was 99.82%.The method was used to detect the viral load in the body of naturally infected iridovirus large yellow croaker,and the results showed that the virus load in the spleen,head kidney and liver of the major immune organs were 8.90×10~4copies/ng DNA,1.86×10~4copies/ng DNA and1.32×10~4copies/ng DNA,respectively.(2)Healthy large yellow croaker was infected by intraperitoneal injection of iridovirus suspension,specific bands of the expected size were amplified by iridovirus PCR after 5d infection(PCR positive detection rate 100%after 10 days of infection)and histopathology observed the characteristic pathologic changes of iridovirus infection-enlarged cells and typical virions,indicating that the population of large yellow croakers artificially infected with iridovirus had been successfully obtained.(3)The mortality rate of artificially infected iridovirus large yellow croaker under a week of continuous starvation and feeding was calculated,the results showed that the mortality rate of large yellow croaker in group A infected with starvation was 14.3%lower than that in group B infected with normal feeding.Starvation after 24 h,liver,spleen,head kidney viral load of starvation group were significantly higher than that of feeding group,after 48h,the viral load of starvation group decreased to the minimum and significantly lower than that of feeding group.Subsequently,the overall viral load in the two groups showed a gradually increasing trend,however,the viral load in the head kidney,spleen and liver of the starvation group was always lower than that of the normal feeding group.(4)Compared with the control group(group C and group D),IFNc,Mx and IgM in spleen and head kidney of starvation group and feeding group were up-regulated to different degrees,and showed a trend of increasing first and then decreasing.Among them,The expression of IFNc in the head kidney and spleen of the starvation group increased more,showing stronger immune stress,the expression of Mx in the head kidney was also significantly higher than that in the feeding group,but it fluctuated over time in the spleen,the expression level of Mx in the spleen of the feeding group was significantly higher than that of the starvation group at the 2nd day,in the rest of the time,the expression level of Mx in the spleen was no less than that of the feeding group,suggesting that starvation helped to improve the expression level of antiviral factors in the body.IgM expression in the two treatment groups showed irregular changes,On the 1st day after treatment,IgM expression levels in the head kidney and spleen of the starvation group were significantly higher than that of the feeding group,which was reversed from the 2nd day to the 3rd day,IgM in the feeding group was higher than that of the starvation group,and this trend remained.(5)Starvation had no significant effect on SOD activity in liver,intestinal,gill and ACP,AKP activity in intestines of artificially infected large yellow croaker.The liver,intestinal and gill CAT activity of the starvation group were all higher than that of the feeding group on the 7th day,and the liver showed significant differences.The ACP and AKP activity of liver,gill in starvation group peaked in starvation after 24 h,and were significantly higher than feeding group.The above results showed that after a period of starvation,the artificial infection large yellow croaker in this experiment had a brief decrease in the viral load,while its immunity and disease resistance were improved in short term(1-2d),preliminary results showed that starvation could slow down the proliferation of iridovirus and enhance the immunity,However,it is not significant under experimental conditions,which is different from the actual application effect of starvation therapy in breeding production.More experiments are needed to further clarify the effect and influence of starvation treatment on the iridovirus disease in large yellow croaker. |
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