Attempts to eradicate graft-transmissible infections through somatic embryogenesis in Citrus ssp. and analysis of genetic stability of regenerated plants

Abstract

Stigma/style somatic embryogenesis (SE) is proved to be effective in the complete elimination of the main Citrus virus and virus-like diseases in 100 % of analysed cases. In the present research SE was applied on 13 genotypes, belonging to the Algerian germplasm collection of two different Citrus species (Citrus limon and C. sinensis) infected by one or more graft-transmissible agents, to evaluate the genetic stability and the sanitation of regenerants. The infected genotypes were regenerated through stigma/style SE and the obtained plants were tested by serological, molecular and biological assays, 6 and 18 months after plantlet grafting, for assessing the elimination of the virus, viroid and virus-like agents present in the mother plants. No evidence of infections of virus and virus–like agents was present in the regenerated plants; whereas, after 18 months up to 24 % of tested plants were found to be infected by viroids, independently from the species and genotypes of the mother plants. Hop stunt viroid (HSVd) proved to be the most infectious viroid; conversely, Citrus exocortis viroid (CEVd) and Citrus viroid III (CVd-III) were present only in ‘Secile’ lemon, as mixed infection with HSVd. SE from stigma/style explants was 100 % effective in the elimination of mixed viroid infections in two lemons (‘Lunario’ and ‘Sans pepins’) and one orange (‘Mitidja navel’). To evaluate genetic stability of regenerants, DNA analyses were performed. No somaclonal variability was observed in lemon regenerants. However, the amplification products of ‘Washington navel 251’ (C. sinensis) revealed genetic instability in some of the regenerated plants.