| Gonadal differentiation in fish is influenced by both genetic and environmental factors.A large number of studies have shown that estrogen complexes including estradiol(17β-estradiol,E2)are widely distributed in the aquatic environment and negative effects on the reproductive system of many aquatic animals including fish.Little yellow croaker(Larimichthys polyactis,L.polyactis)is one of the important economic marine fish in China,Korea and Japan.However,due to overfishing,seawater pollution and other factors,the survival of L.polyactis population is seriously threatened.To investigate the effects of estrogen on the reproductive system of L.polyactis,so as to better protect the L.polyactis population,histological identification of the process of gonadal development and sex differentiation in L.polyactis was firstly carried out in the present study.The results showed that the ovary development was relatively continuous,but in male gonad development,we discovered a 37-days transient hermaphroditic stage during the period of43-80 dph(days post-hatching,dph),which was characterized by the presence of early primary oocytes(EPOs)in testes,followed by apoptosis and degeneration.To explore the molecular mechanism of this phenomenon,estrogen E2,androgen 11-ketotestosterone(11-KT)and Testosterone(T)level were measured at a total of 15 time points of 50-720 dph in L.polyactis.The results showed that this transient hermaphroditic stage is caused by the absence of 11-KT and elevated levels of E2,and 11-ketotestosterone(11-KT),not testosterone(T),is the major androgens in L.polyactis.To further prove that the role of E2 in the transient male hermaphroditic stage,the present study carried out E2 exposure experiment on L.polyactis from 30 dph which was start to sex differentiation at histological level.The L.polyactis were exposed to E2(10 μg/L)from 30-90 dph,and after exposured,the E2 withdrawal and continual cultured to 150 dph.Histological results showed that E2 exposure caused three major phenotypes at 30 and 60 days after treatment(dat),including ovary,ovotestis and gonadal development retardation compared with the control groups.The average ratio of these three phenotypes is 60.6%,11.97% and 27.43%,respectively.Meanwhile,E2 expose inhibited the growth of L.polyactis,which mainly demonstrated that body length and body weight were significantly lower than the control groups.However,after E2 withdrawal 60 days,the body length and body weight were recovered,but the phenotype of male gonadal development retardation was still not recovered.To further explore the molecular regulation mechanism of major sex-related genes after E2 exposure in L.polyactis,the expression of these genes was studied by q RT-PCR.The results showed that compared with the wild-type group with male gonadal phenotype,the expression levels of female related genes cyp19a1 a and foxl2 were significantly increased in the E2 exprouse groups,while the expression levels of male related gene dmrt1 were significantly decreased in the ovotestis gonads group,while the expression levels of figla gene were not significantly changed.In conclusion,E2 is a necessary but not sufficient condition for the formation of ovotestis gonads in L.polyactis,and the molecular regulation mechanism of E2 inducing sex reversal to female in L.polyactis still needs to be further studied.This study not only enriched the reproductive biology of fish and the molecular mechanism of sex determination and differentiation,but also provided basic data for the theory of sex control of L.polyactis,and had positive significance for the protection of the ecological population of L.polyactis. |
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