| Orchids often have a deep symbiosis relationship with other organisms such as fungi, and mycorrhizal fungi are their live source of nutrition. In fact, mycorrhiza is not just a symbiosis between fungi and plants, but there are endophytic bacterias. Endophytic bacterias play an important role in the life history of orchids, they can effect the formation of mycorrhiza and its stability of plant-fungi interaction. We isolated endophytic bacterias from the roots of wild Dendrobium catenatum, which is called "plant gold". Separately, inoculated with the seeds and seedling of D. catenatum to screen the benefical endophytic bacterias, then identified them to enrich the resources of orchids endophytic bacterias. Simultaneously, in order to investigate growth-promoting mechanism of those strains, we tested their plant growth promoting activities(PGPA). Then, combing 8 strains of endophytic fungi (C20, C22, C35, C36 and C42 were isolated from wild D. catenatum; L12, L28 and L24b from wild D.Loddigesii), seedlings of D. catenatum were cultured together with endophytic bacterias and fungi after non-antagonistic screening, to study the effect of endophytic bacterias-fungi interaction on the growth of D. catenatum. The main results were as follows:(1) 19 endophytic bacterias were isolated from the roots of wild D. catenatum using culture-dependent approaches, and we got 6 growth-promoting strains by inoculating with the seeds and seedlings of D. catenatum, respectively. Among the 6 strains,4 strains (TX-2, TX-5, TX-7 and TX-19) had positive effect on seed germination and protocorm growth; 5 strains(TX-5、TX-7、TX-12、TX-16 and TX-19)could positively promote the growth of seedlings.(2) According to morphological characters and 16S rDNA sequence analysis, the 6 strains mentioned above were attributed to Bacilllus megaterium(TX-2), Ochrobactrum(TX-5), Erwinia(TX-12), Enterobacter(TX-7, TX-16), and Herbaspirillum(TX-19). Whereas, Ochrobactrum was isolated for the first time from orchids, and the highest similarity of TX-7, TX-16 were 95.6% and 96.1%, they may be potential novel taxonomic units.(3) PGPA analyses showed that 6 strains mentioned above had different levels in fixxing nitrogen, solubilizing phosphate, secreting IAA and producing siderophore. Among them, TX-2 could fix nitrogen and secrete IAA, TX-5 could secrete IAA and produce siderophore, TX-7 and TX-16 just couldn’t fix nitrogen. Whereas, TX-19 had all the biological characteristics.(4) Through antagonistic experiments, there were 23 antagonistic combinations and 25 non-antagonistic ones. By co-culturing above 25 combinations with seedlings of D. catenatum for 120 d,6 combinations were screened step by step with positive effect on the weight net growth rate, then further determined the number of new roots and shoots, and root branching degree for variance analysis, results showed C22+TX-19, L28+TX-16 and L28+TX-19 were advantage combinations, which all induced a significant enhancement in some norms of seedling compared with singly inoculation, and they had synergistic effect or additive effect in seedlings of D. catenatum.(5) Through re-isolating experiments, we found that, the homologous strains were all re-isolated from the controlleds in addition to seedlings noculated with C36, and no strain was re-isolated from the controlled. It showed that single or mixed microorganisms all established a symbiotic relationship with seedlings of D. catenatum bating C36. While, TX-5 and TX-7 were re-isolated from the seedling, this suggested they were beneficial for C36 to infect the roots of D. catenatum.(6) As a result, only 3 advantage combinations were screened out from 48 groups of "fungi + bacteria", it showed bacterial promotion on seedling of D. catenatum to be fungal specific to some extent. But TX-19 could create a positive interaction effect on the growth of seedling of D. catenatum combinated with 2 different fungi (C22、 L28)from 2 different orchids. Therefore, we speculated that bacterial promotion has a low degree of fungal specific and plant specific. |
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