The Antibacterial Effects Of Resveratrol On Pathogenic Aeromonas Hydrophila

Resveratrol(RES)is a non-flavonoid plant polyphenol,whose chemical name is 3,5,4’-trihydroxysti lbene.Resveratrol is named for its first separation from the roots of resveratrol.Resveratrol is a substance secreted by the plant for antibacterial and anti-oxidation which are stimulated by pathogen invasion,ultraviolet radiation and ozone.Aeromonas hydrophila is a widely distributed gram-negative bacterium in the freshwater ecological environment,which is ubiquitous in the natural water,soil,aquatic animal body surface and digestive tract.A.hydrophila can cause local or systemic infection of aquatic animals under specific conditions.Fistly,this research analyzed the inhibitory effects of resveratrol on the growth,biofilm formation and 他 the activity of hemolytic enzyme of A.hydrophila in vitro,as well as the effects on the expression of virulence factors and gene in the sensory regulatory system of A.hydrophila.Secondly,the effects of resveratrol on the growth and antioxidant capacity of Grass carp kidney cell(CIK)cells was exmained by using the extracellular product of resveratrol co-cultured with A.hydrophila.Artificial intraperitoneal injection of A.hydrophila into Carassius aurat.us gibelio to detect the changes of antioxidant capacity in liver,plasma biochemical indexes and the expression of genes related to inflammation.Finally,lipopolysaccharide(LPS)was used to induce immune stress in CIK cells to explore the effects of resveratrol on growth and antioxidant capacity of CIK cells in this study.The expression of antioxidant and inflammatory genes in CIK cells was detected by reverse transcription-polymerase chain reaction(RT-PCR).1.The minimal inhibitory concentration(MIC)of resveratrol against A.hydrophila was above 1024 μg/mL,and the concentration of resveratrol<64 μg/mL did not affect the growth ratio of A.hydrophila.When the concentration of resveratrol ≥ 32 μg/mL,could significantly inhibit the biofilm formation and the activity of hemolytic enzyme of A.hydrophila(P<0.05),which were dose-dependent.RT-PCR analysis showed that resveratrol could significantly up-regulate the expression of luxR mRNA in the sensory regulatory system of A.hydrophila(P<0.05),and the expression of luxS mRNA was down-regulated significantly(P<0.05),while the expression of omp mRNA decreased significantly(P<0.05).2.When the concentration of resveratrol was≤4 μg/mL,there was no significant effect on the growth activity of CIK cells(P>0.05).When the concentration of resveratrol≥ 16 g/mL was co-cultured with NJ-35,bacterial extracellular products had no significant effects on the growth activity of CIK(P>0.05)and the concentration of LDH in cell culture medium was significantly decreased(P<0.05).Resveratrol could enhance the activity of SOD and the content of GSH and reduce the content of MDA in CIK cells,which indicated that the toxicity of the extracellular product was weakened and the damage to CIK was reduced.The mortality of Carassius auratus gibelio with intraperitoneal injection of resveratrol with 25 mg/kg,50 mg/kg and 100 mg/kg before 1 h A.hydrophila challenged was significantly decreased(P<0.05).Meanwhile,compared to the challenged group,the concentration of plasma AST and ALT were significantly decreased(P<0.05)and the concentration of plasma ALP and ALB was significantly increased(P<0.05),as well as the content of MDA in liver significantly decreased(P<0.05)and the activity of CAT changed inconspiciouly(P>0.05)in the injection of 25 mg/kg and 50 mg/kg resveratrol groups.Among them,the activity of SOD in liver in the injection of 25 mg/kg resveratrol group was significantly higher than that in the challenged group(P<0.05).Compared to the challenged group,the concentration of plasma AST and the activity of SOD in liver in the injection of 100 mg/kg resveratrol group changed inconspiciouly(P>0.05).The concentration of plasma ALT decreased significantly and the concentration of plasma ALP and ALB significantly increased(P<0.05),as well as the content of MDA in liver significantly decreased and the activity of CAT increased significantly(P<0.05)in the injection of 100 mg/kg group.RT-PCR analysis showed that the expression of TNF-αand IFN-y mRNA significantly decraesed in all resveratrol treatment groups(P<0.05).The expression of IL-10 mRNA was significantly down-regulated in the injection of 50 mg/kg group.3.The activity of CIK cells significantly decreased and the concentration of LDH in cell culture medium significantly increased by treated with LPS(P<0.05).When the concentration of resveratrol was 2 μg/mL in the culture medium,compared to the LPS treatment group,the activity of CAT and the content of GSH significantly in liver increased and the content of MDA significantly decreased(P<0.05),while the expression of CAT mRNA significantly increased and the expression of TNF-α mRNA significantly decreased(P<0.05).When the concentration of resveratrol was 4 μg/mL in the culture medium,compared to the LPS treatment group,the activities of CAT and SOD in liver significantly increased and the concent of MDA significantly decreased(P<0.05),while the expression of CAT and GSTA mRNA increased and the expression of IL-1,IFN-y and TNF-α mRNA significantly decreased(P<0.05).