Ascospore germination characteristics and the ultrastructure observation of fruit body of Pseudopeziza medicaginis

Abstract:

[Background] Medicago sativa common leaves spot caused by Pseudopeziza medicaginis is a main fungal disease in M. sativa. This strain can overwinter by the formation of apothecia and then release ascospore to complete primary infection. Thus we observed the fruit body under an electron microscope for exploring the colonization mechanism of P. medicaginis. [Objective] To clarify the conditions of ascospore germination, whether the Ca2+ signal pathway is involved in the form of appressoria and observe the ultrastructure of the ascospore and fruit body of P. medicaginis. [Methods] Effect of different types of carbon and nitrogen sources and the different doses of Ca2+ on ascospore germination and appressoria form were studied by optical microscope. The ultrastructure of the fruit body and ascospore were observed by a transmission electron microscope. [Results] Carbon and nitrogen sources benefited the ascospore germination. Glucose, maltose, sucrose, amino acetic acid, yeast powder, urea, peptone, ammonium nitrate, sodium nitrate and so on could induce the formation of appressorium. Urea had the strongest effect. The Ca2+ not only could promoted the germination of ascospores but also could induce the formation of appressorium, the effect was better than adding carbon and nitrogen sources. With Ca2+ concentration increased, the rate of ascospore germination and the rate of appressoria formation increased, the best dose of Ca2+ was 10 mmol/L, once exceed this dose, the rate of ascospore germination and the rate of appressoria formation decreased. With exogenous Ca2+ or A23187 which is Ca2+ carrier addition, the inhibition role of EGTA could partially relieved. Neomycin, an inhibitor of phospholipase, nicardipine which could block Ca2+ channels, chlorpromazine, CAM antagonist could inhibit the germination of ascospores of this strain. The mature P. medicaginis fruit body broke through the epidermis of the host, and the middle part of the fruit body cracked to expose the ascus. The surface of ascospore had a ring-shaped pattern, the nucleus was dark, the number of organelles such as mitochondria is large, the number of lipids was large and developed, and the cells were filled with high electron density materials. [Conclusion] Ca2+ signal pathway was involved in the process of ascospore germination and charge of the appressoria formation of P. medicaginis.