Screening Of Natural Drugs Against Cryptocaryon Irrrtans And Study Of Cryptocaryon Irrrtans Theront Apoptosis Induced By Nerolidol

Cryptocaryon irritans parasitizes marine fish and causes fatal marine fish“white spot disease”.The disease has seriously hindered the development of aquaculture industry in southeast China.Therefore,it is urgent to find safe and effective control methods.Chinese herbs have the advantages of wide source,low toxicity,low drug residue,no pollution and so on.Moreover,Chinese herbs contain a variety of phytochemicals and has a variety of pharmacological effects,which provides a large number of drug sources for screening new drugs against cryptocaryonosis.Therefore,in this paper,10 Chinese herbs and 14phytochemicals were screened out to test the inhibition on Cryptocaryon irritans in vitro.The control effect of the most effective phytochemicals on the cryptocaryonosis of Trachinotus ovatus was tested;study its effect on innate immune function of Trachinotus ovatus;the concentration range of inducing and stimulating the apoptosis of Cryptocaryon irritans theronts was preliminarily determined,and the main results are as follows:1.The inhibition effects of 10 Chinese herbs on Cryptocaryon irritants in vitro:The results showed that the ethanol extracts of Agrimonia pilosa,Zanthoxylum,Catechu,Polygonum Aviculare and the water extract of Catechu caused 100%mortality on Cryptocaryon irritants theronts within 30 min.And the ethanol extracts of Polygonum Multiflorum,Spina Gleditsiae,Celosia cristata,Apricot kernels,Verbena officinalis and the water extract of Agrimonia pilosa had significant killing effect on Cryptocaryon irritants theronts.In addition,the water extract of Catechu 100%inhibited the hatching of Cryptocaryon irritants tomonts within 7 days.2.The inhibition effects of 14 phytochemicals on Cryptocaryon irritants in vitro:The results showed that nerolidol,citronellal,menthone,linalool,geranyl acetate,1,8-cineole,geraniol,citral,and thymol from the 14 phytochemicals had significant killing effect on theronts.And nerolidol possessed the highest inhibitory effect on Cryptocaryon irritants theronts.The half-maximal inhibitory concentrations(IC₅₀s)of nerolidol on Cryptocaryon irritants theronts,protomonts,and tomonts were 2.1×10^-3,1.33×10^-2,and 7.1×10^-3μL/mL,respectively.At 1.0×10^-2μL/mL,nerolidol significantly changed the morphology of Cryptocaryon irritants theronts,and the somatic cell membrane of theronts was broken.At3.0×10^-2μL/mL,nerolidol made the color of Cryptocaryon irritants protomonts lightened and protomonts created bubbles around their membranes.At 1.0×10^-2μL/mL,the cytoplasm of the Cryptocaryon irritants tomonts treated with nerolidol shrank significantly.3.The efficacy of nerolidol on cryptocaryonosis of Trachinotus ovatus:The results showed that a diet supplemented with 2.3 mL/kg of nerolidol reduced the number of Cryptocaryon irritants trophonts on pompano gills and the chemotaxis of Cryptocaryon irritants theronts to pompano mucus and sera and did not cause inflammatory reaction in vivo and increased the cumulative survival rate of Trachinotus ovatus.4.Effects of nerolidol on non-specific immune function of Trachinotus ovatus:The results showed that a diet supplemented with 2.3 mL/kg of nerolidol caused the activity of superoxide dismutase（SOD）and catalase（CAT）increased,the activity of acid phosphatase（ACP）decreased,and the concentration of total protein（TP）increased.This suggested that nerolidol improved the total protein of Trachinotus ovatus,reducing free radical production and lipid damage,thereby promoting fish health.5.The range of concentrations of nerolidol induced apoptosis of Cryptocaryon irritants theronts:the concentration of inducing and stimulating the apoptosis of Cryptocaryon irritants theronts ranged from 2.0×10^-3μL/mL to 6.0×10^-3μL/mL.When the concentration of nerolidol was 2.0×10^-3μL/mL,the thronts apoptosis began.When the concentration of nerolidol was 4.0×10^-3μL/mL,the theronts apoptosis was the highest,and the theronts necrosis began at 6.0×10^-3μL/mL.The results showed that nerolidol had a strong inhibitory effect on the Cryptocaryon irritants in vitro and in vivo.In addition,nerolidol improved the total protein of Trachinotus ovatus,reducing free radical production and lipid damage,thereby promoting fish health.The concentrations of nerolidol induced and stimulated the apoptosis of Cryptocaryon irritans theronts ranged from 2.0 to 6.0×10^-3μL/mL.In conclusion,nerolidol has great development prospects in the control of marine fish cryptocaryonosis.