Effects Of Two Main Earthy Compounds On Immune Function Of Zebrafish

Dirty smell is a bad smell in the body of fish.It is a general term for a variety of compounds.The main ingredients are 2-methylisobome(2-methylisobome,2-MIB)and Geosmin.These two terpenoids are mainly secondary metabolites produced by actinomycetes and freshwater planktonic algae.During the flourishing period of the growth of these microorganisms,the fishy smell also shows an increasing trend and reaches the peak.At present,the synthesis pathways of 2-MIB and Geosmin in actinomycetes and cyanobacteria are well defined.2-MIB is synthesized by double genes,while Geosmin is formed by a single bifunctional enzyme.They are mainly through the gills of fish or feeding into the fish and accumulate in the body,making the fish produce an unpleasant smell of earth.The odor can be detected in all water bodies,such as ponds,lakes and rivers,but there are few reports on the effect of the odor on the organisms living in this water environment.The effects of 2-MIB on fish other than causing the fishy smell are unknown.Recent studies have found that Geosmin has certain effects on the physiological function of fish,but the specific mechanism still needs to be further clarified.In order to investigate whether 2-MIB and Geosmin have effects on immune function of fish,the 3-month-old zebrafish was exposed to 2-MIB at a concentration of42 ng/L and Geosmin at a concentration of 10 ng/L,and the gills tissues of zebrafish were taken for trantrantome sequencing analysis.A total of 728 differentially expressed genes were found in the 2-MIB experiment transcriptome(163 genes were up-regulated and 565 genes were down-regulated),and 121 KEGG pathways were enriched in the differentially expressed genes(29 up-regulated and 92 down-regulated),among which 7immune-related KEGG pathways were screened out,consisting of a total of 23 genes.Transcriptome analysis of the Geosmin experiment revealed a total of 1,944 differentially expressed genes(527 genes were up-regulated,1417 genes were down-regulated),and 215 KEGG pathways were enriched by the differentially expressed genes(92 up-regulated,123 down-regulated),among which 8 KEGG pathways were mainly related to immunity,and a total of 69 genes were found.At the same time,the two Transcriptome had a total of 507 same in the differential genes.Some differentially expressed genes obtained from the two experiments were enriched into the Oxidativephosphorylation pathway,and the expressions of several genes in this pathway were up-regulated.In down-regulated KEGG genes,the expressions of Toll-like receptor signaling pathway,NOD receptor signaling pathway,Salmonella infection,RIG-I-like receptor signaling pathway,Adipocytokine signaling pathway,JAK-STAT signaling pathway and p53 signaling pathway,which are mainly related to immune pathways,were all reduced.In order to further verify the effect of 2-MIB on Oxidative phosphorylation and immune-related gene expression in zebrafish,the expression of genes in oxidative phosphorylation pathway and Toll-like receptor signaling pathway was verified by fluorescence quantitative PCR,and the change of superoxide dismutase SOD activity was further verified.The results indicated that 2-MIB significantly upregulated the expression of oxidative phosphorylation related genes ndufb7、mt-cyb、mt-nd4、mt-nd6、mt-co2 and mt-atp6,while Toll-like receptor signaling pathway related immune genes rela、cd40、ikbkb、mapk8b、mapk3 and ripk1 l were significantly down-regulated.SOD activity of superoxide dismutase increased in gill and liver tissues after 2-MIB exposure.The results indicated that 2-MIB in water could increase the level of free radicals in fish,cause cell damage,and possibly lead to the decline of related immune function.In order to explore the effect of Geosmin on the immune function of zebrafish,RT-q PCR was used to verify the expression of immune-related genes in gill,liver,kidney and head kidney of zebrafish treated with Geosmin,and to further determine the changes of reactive oxygen species ROS and nitric oxide NO in the control group and the Geosmin treated group.The results showed that Geosmin significantly down-regulated the expression of immune-related genes irf4a、casp7、cd40、il26、irg1and nos2 a.After Geosmin exposure,the content of ROS and NO decreased in zebrafish.These results indicate that Geosmin in water may decrease the resistance to microorganisms and the bactericidal activity of cytokines in fish,and may decrease the related immune performance.The fishy smell of fish not only causes the economic loss of aquaculture,but also reduces the desire of consumers to buy.Although,the smell has not affected human health,the smell in water has become a common complaint of drinking water consumers.Therefore,it is of practical significance to explore the potential effects of two main earth-smelling compounds,2-MIB and Geosmin,on zebrafish.At the same time,it has important implications fos aquatic life and aquaculture.