Spore Germination And Dual Culture Of Arbuscular Mycorrhizal Fungi

Arbuscular mycorrhizae can enhance the ability of absorbing P in the soil for plants, improve the nutrition level in plants and increase the assimilating area of roots and subsequent tolerance to drought. This is of particular interest in plant such as citrus tree with few hairy roots leading to the P uptake difficulty. So it is significant to produce massive arbuscular mycorrhizal fungi propagates for application on citrus planting to increase fruit output and improve fruit quality. In the present paper, four representative propagates of arbuscular mycorrhizal fungi including spores of Gigaspora margarita, Glomus mosseae and Glomus sp as well as sporocarps Glomus sp were used to investigate the influence of low temperature storage and different propogates on germination in vitro culture. On the other hand, the infection mechanism of hyphae to host plant roots in dual culture was also probed. The main results were as follows:1. A lot of indigenous spores, which mostly belong to Glomus were separated from soil around rootsphere of citrus trees, which provided material for the studies on influence of different time chilling storage on the germination of indigenous spores.2. The structures of arbuscular mycorrhizae in citrus roots was observed, in which there were some typical mycorrhizal structures, such as hyphae, vesicule and arbuscule. AM fungi established favorable symbiosis relation with citrus roots.3.Indigenous spores isolated from soil were stored at 4C for 0, 10, 20, 30, 40, 50 days respectively. Then they were cultured on the plates with 0.8% water-agar medium in the dark at 25 C before the germinating spores were counted. The results showed that storage of spores at 4C for 10~50 days dramatically enhanced the spore germination percentage; The best results came from 20 days' treatment, in which the spore germination percentage was 30.34% whereas the spores without chilling storage germinated few, only 4.94%.4.The comparison of germination percentage of different propaga'tes between intro-and inter-species of arbuscular mycorrhizal fungi in the same culture condition was made. The highest germination percentage is Gl sp sporacarps, amount to 60%; while the lowest is Gl. sp spores, only 7.14%. The difference of germination percentages among Gi. margarita, Gl. mosseae, Gl. sp sporacarps and Gl. sp single spore was significant.5. The clover roots were shaky cultured in improved White liquid medium, from which the in vitro grown roots were obtained for dual culture. The dual culture system ofGi. margarita with clover roots were established. New spores were observed in the culture system after the germinated spores co-cultured with roots in vitro for one month.6. The studies on mechanism of infection and sporulation showed: 1) one or several hyphal stretched from handle cell and then branched gradually. 2) Clover roots had no prominent influence on the germination of spores. 3) Two weeks later, the branches in dual culture system were much more than control in which there were no clover root segments co-cultured with AM fungi spores. The phenomenon of reptant infection to host roots was observed. 4) Three weeks later, some runner hyphae stopped elongation and formed a ball at the tip, under which ramified some septal hyphae, among which some formed branched absorbing structure (BAS). 5) After four weeks, hyphae spread all over petri dishe plates. At the same time, arbuscular-like hyphae extending from roots were observed nearby whereas the hyphal growth speed in the control slowed down and even stopped. 6) Polarity experiment showed that ramified hyphae had geotropism character. 7) The observation result of clover roots dyed with trypan blue in dual culture showed that roots were infected efficiently by AM fungi. There were many new spores in the cortex of roots, but no one in the control.