Growth Promotion, Stress Tolerance And Disease Resistance In Rice Conferred By Piriformospora Indica And The Preliminary Study Of Mechanisms

Piriformospora indica, a root endophytic fungus, which was isolated from Thar Desert in India, exhibits symbiotic association with a variety of host plants. This fungus imparts a broad spectrum of benefits to its host, such as growth-promotion, disease resistance, and high salt tolerance.Rice is the main food crop for nearly half of the global population. However, the cultivation of rice has been always affected by various biotic and abiotic stresses, and further growth in rice production is limited. In the current agricultural production, people mainly use chemical fertilizer, pesticide and other methods to promote growth and improve yields. But unreasonable application of fertilizers and pesticides often cause soil compaction, environmental pollution, increased pesticide residues and other environmental problems. Promoting the growth of rice, making rice adapt to changes in the environment, improving the yield and quality of rice, achieving sustainable development of agriculture industrial by biological pathways are directions of research in recent years. The thesis is mainly about growth promotion, stress tolerance, and disease resistance in rice conferred by P. indica and the preliminary study of their mechanisms. The salient findings of the research are:1. Established the co-cultivation systemto study the effect of P. indica in rice.We found when5mycelium blocks of P. indica and6rice seedlings(4days old) were co-cultured on two different medium-MS and PNM, P. indica can faster contact with the rice roots on PNM medium, and showed significant growth-promoting effect. Microscopic analyses revealed that chlamydospores of P. indica were detected in intracellular and extracellular of rice root cortex at14th day after co-cultivation.2. P. indica promoted the growth of aerial part in rice, and enhanced rice yield.After rice seedlings were co-cultured with P. indica, the height, biomass of shoot, activity of root and nitrate reductase activity of rice colonized by P. indica were significantly higher than that in un-colonized one. The chlorophyll content in treated rice increased by19.77%and17.89%respectively after P. indica treatment for3and4weeks. Compared with the control, the number of tillers increased by10%-20%, and the number of per panicle and filled grains increased by more than20%in rice colonized by P. indica. The colonized rice headed4-6days earlier than the un-colonized ones. Results of the RT-PCR analysis showed that the expression of auxin-regulated genes OsIAAlS and YUCCA were up-regulated in leaves of rice colonized by P. indica, and the expression levels increased by49.5%and30.4%respectively. The expression of NRR, which negatively regulates the growth of rice, decreased by41.9%as compared to control.3. P. indica enhanced the tolerance of rice to cadmium stress, and significantly decreased the accumulation of cadmium in shoot of rice.P. indica can promote the growth of rice seedlings under cadmium(Cd) stress, including improvement of biomass accumulation, increase of the height and root length of rice plants. Under low levels Cd stress (3mg/L and10mg/L), neither the plants colonized by P. indica or not exhibit symptoms of Cd poisoning in12d after treatment. Under25mg/L and50mg/L Cd stress, the plants colonized by P. indica exhibited symptoms of Cd poisoning at the12th day and the10th day, and were2d and3d later than control respectively. By measuring chlorophyll content and malondialdehyde (MDA) content, we found P. indica can increase chlorophyll content and decrease MDA content in plants, accordingly alleviating Cd toxicity to rice, improving rice tolerance to Cd stress. By measuring the Cd content, we found that Cd content in the root of variety Yueyou9113seedlings colonized by P. indica was9.84folds of that in control, while decreased by38.4%in the shoot. At tillering stage and filling stage, Cd content in the root of treated plants was increased24.1%compared with the control, but Cd content in different tissues of shoot was decreased from33.5%to48.0%. For variety Qianyou2, at tillering stage, the Cd content in colonized plants was decreased from45.5%to73.6%as compared to control. And at filling stage, the Cd content in root, leaves and unpolished rice of treated rice were decreased15.8%,25.9%and53.6%respectively. The results of RT-PCR analysis indicated that the expression of OsHMA3, which has been identified as a regulator for xylem Cd transport in rice by mediating vacuolar sequestration of Cd in root cells, was up-regulated obviously as compared to control, meanwhile the expression of OsHMA2, an efflux-type metal transporter, was down-regulated. These changes induced by P. indica may reduce migration of Cd from root to shoot in rice.4. P. indica strengthens salt-stress tolerance in riceSalt stress experimental results showed that, P. indica promoted the growth of rice seedlings under salt stress, and enhanced tolerance to salt stress in rice. After growing7d on PNM medium with100mM/L NaCl, the height and root length of rice seedlings conferred by P. indica increased18.6%and27.7%respectively, and the fresh weight and dry weight respectively increased35.3%and16.9%compared to control. After watering the potted rice seedlings with solution of200mM/L NaCl, the degree of damage was significantly alleviated in rice colonized by P. indica in comparison with the control plants at the14th day and the21th day. After salt treatment for a month, nearly half of the rice seedlings in the control group had died, and the withered rate of plants colonized by P. indica was reduced by20%.5. P. indica increases resistance to rice sheath blight、bacterial leaf streak、bacterial leaf blight.The Rhizoctonia solani, Xanthomonas oryzae pv. oryzacola and Xanthomanas oryzae pv. oryzal were inoculated on rice which had co-cultivated with P. indica for14d. The results showed P. indica enhanced resistance of rice to sheath blight, bacterial leaf streak and bacterial leaf blight. At the2th day after inoculating R solani, the plants in the control group appeared brown lesions, and lesions had coveraged stem at the5th day. Large moire lesions appeared on the leaves of the control group at the7th day. Brown lesions appeared in the plants colonized by P. indica at the4th day, and the lesions were limited to the stem at the7th day. At the6th day after inoculating X. oryzae pv. oryzacola by by the method of leaf-needling, water-soaked lesions appeared in the control group, but lesions were limited to the inoculation sites in the treatment group. The length of lesions in the treatment group was significantly shorter than control, which is half the length of the lesion compared to control group at the15th day. At the3th day after inoculating X. oryzae pv. oryzal, inoculation sites in the control group appeared yellow withering, while treatment group were scarcely illness. The control and treatment group both appeared symptoms of yellow withering, green withering, and leaf curl at the8th day. However, the expansion of the lesions in treatment rice were limited, and the length of lesions was decreased by47.3%compared to control. The relative immunization efficiency to bacterial leaf blight in rice colonized by P. indica reached to40.68%. P. indica enhanced resistance to bacterial leaf blight in rice, by increasing proline content and polyphenol oxidase activity in rice leaf, inducing the expression of disease resistance-related genes PAL, OsBTF3, Rcht2and PR1α.