Morphogenesis of protoplasts of Entomophthora egressa in simplified culture media

Abstract

Protoplasts of Entomophthora egressa MacLeod and Tyrrell were grown in Grace's medium and in two simplified liquid media. These latter media contained reduced numbers of amino acids as compared with Grace's insect tissue culture medium. The pattern of protoplast regeneration was the same in all three media used, but additional stages in the morphogenetic sequence werefound. A spherical mesoprotoplast and an elliptical mesoprotoplast stage with actively moving filopodia were detected early in the growth cycle. The clumping of rod-shaped and spherical hyphal bodies was observed. Mycelial development was very pronounced by 89 h of incubation. The osmolarity and pH of the media were unchanged before this time. The individual hyphae were predominantly initiated by spherical hyphal bodies.All stages of regeneration produced spindle-shaped protoplasts when added to fresh media. The inoculation of spent media with spindle-shaped protoplasts favoured an accelerated morpho-genesis of the cells through the presently reported stages.