Identification and partial characterization of Olyra longicaudata (McClelland, 1842) vitellogenins: Seasonal variation in plasma, relative to estradiol-17β and ovarian growth

Abstract

This study aims at immunochemical characterization of plasma vitellogenin (VTG), development of an heterlogous VTG ELISA and to relate seasonal variation in plasma VTG and estradiol-17β (E2) levels with ovarian growth (gonadosomatic index, GSI) in Olyra longicaudata (McClelland, 1842), a rare hill-stream catfish endemic to North East India. On native PAGE, plasma from E2-injected male, vitellogenic as well as gravid females, but not untreated male, resolved into two major protein bands. These two proteins stained positive for carbohydrate, lipid and phosphorous, albeit with differential intensity and cross-reacted well with catfish VTG antiserum (a-VTG) suggesting them as putative VTGs in circulation. Ammonium sulphate (50%) fractionation followed by SDS-PAGE analysis of E2-treated male plasma resolved into four protein bands (150–15kDa), of which two, with molecular mass of 150 and 130kDa cross-reacted with a-VTG indicating them as VTG monomers. Immunoprecipitation of E2-induced plasma and immunoblot analysis of crude yolk proteins with a-VTG revealed two proteins in each case indicating two forms of VTG, present in circulation, possibly act as yolk precursors. Competitive antigen-capture ELISA developed earlier for catfish, Clarias batrachus VTG (CF-VTG1), revealed parallel binding slopes between dilution curves of plasma from vitellogenic female, E2-treated male and CF-VTG1 standard. Congruent with gradual increase in plasma E2, ovarian weight and appearance of vitellogenic and yolky oocytes, VTG level in circulation increased sharply in May–June, reaching the peak value in July, dropped sharply during August–September and was undetected or negligible in amount during December allowing identification of the ripening, the pre-spawning, the spawning and the quiescent phases respectively.