| Wheat stripe rust (WSR) is an economically important fungal disease caused by Puccinia striiformis west.f.sp.tritici Eriks et Henn on wheat. Featured by wide attack region, high epidemic frequency as well as serious hazard and yield loss, this disease has become a major constraint for wheat production worldwide. The controls of stripe rust are mainly depending on utilization of resistant varieties, chemical fungicides and integrated agronomic measures. Due to variability of races in the pathogen population and chemical residues in the environments, it is necessary to develop other ecofriendly and sustainable means for the control of stripe rust. For unique advantages and diverse biological properties, bacterial endophytes have attracted the attention of many plant pathologists. So far, the information on utilization of bacterial endophytes in controlling stripe rust is too limitted. In the dissertation, the author aims to investigate the populations of bacterial endophytes within wheat plants, and screen the isolates with high growth-promoting, WSR-controlling and yield-enhancing activities, and clarify the mechanisms on the suppression of WSR and the enhancement of yields by the bacterial endophytes. The main results are summarized as follows:1. A total of 313 bacterial endophytes were isolated and their population diversity analyzed from various organs of wheat plants at different growth stages in different regions. Of the strains obtained,101 (32.27%) were isolated from the samples collected in Wolong District in Nanyang city. The isolation qualities of the bacterial endophytes significantly differed from one organ to another, and varied with growth stages. The maximum numbers of bacterial endophytes were 11.5×105cfu/g,6.88×105cfu/g,4.77×105cfu/g and 3.47×105cfu/g at whole growing stage in the samples of roots, stems, leaves and grains, respectively. The numbers of bacterial endophytes isolated from different organs were 2.73×105cfu/g, 6.65×105cfu/g,10.48×105cfu/g and 9.3×105cfu/g at the tiller, enlongation, heading and waxy ripe stages, respectively. Molecular identification of the strains was carried out based on 16S rDNA sequences. These strains were grouped into 23 operational taxonomic units (OTU) including 9 genera based on their maximum identity with those in GenBank detabase. Bacillus was the dominant genus, contributing to 76.37% of the total numbers of bacterial endophytes isolated, followed by Pseudomonas (10.23%). Bacillus cereus and Bacillus subtilis had a higher isolation frequency (9.7% and 4.37%, respectively) in the whole stage. This is the first report of endophtic Stenotrophomonas maltophilia isolated from winter wheat plants.2. A total of 127 bacterial endophytes were further tested. Their growth-promoting activities on wheat seedlings were investigated. The data obtained were evaluated using multiple statistical methods including cluster analysis, discriminatory analysis and comprehensive correlation analysis. Six isolates (SB 127, LD161, RA135, JD204, RC79 and RBI32) had higher efficacies in enhancing leaves length, dry weights of stem, leaf and root, and showed higher IAA-producing, inorganic/organic phosphorus-dissolving activities. A significant positive correlation was found between growth promotion and their IAA productivity or phosphorus solubility. The IAA productivity and inorganic/organic phosphorus-dissolving activities were found to be responsible for wheat growth promotion. The wheat-growth-ptomoting activity significantly varied from one isolate to another, and could be used as indicators for the screening of bacterial endophytes as biocontrol and yield-enhancing agents for sustainable wheat production.3. The strains RC79, SB127, RB132, RA135, LD161 and JD204 were identified as Bacillus cereus, Bacillus megaterium, Bacillus subtilis, Pseudomonas fluorescens and Pseudomonas putida based on their morphological, physiological and biochemical characteristics as well as 16S rDNA sequences and rpoD gene (LD161 and JD204). The optimum growth temperatures for these strains were 28℃～30℃. The best pHs the growth of the strains were 6-8. Glucose could be utilized as a carbon source by strains RC79, SB 127, RB132 and RA135. Both isolates LD161 and JD204 could not use xylitol as a carbon source for their growth. The six strains almost could use all of the 8 nitrogen sources tested. The activity of arginine decarboxylase, lysine decarboxylase were detected among the strains except RB132. The six strains with drug resistance marker could enter into wheat plants by colonization test. The three strains (SB 127, RB132 and JD204) could move to the abovedground organs from the roots of the wheat plants inoculated.4. The efficacies of six bacterial endophytes (RC79M, SB127M, RB132M, RA135M, LD161M and JD204M) in controlling WSR were tested on 14 different wheat cultivars in field trail. The mean disease incidences were 14.62% ～19.9% on the wheat cultivars, which was 68.9%～83.4% lower than their respective controls. The mean disease indices of the plots treated by the bacterial endophytes were 4.98～7.13, which was 64.7%～92.3% lower than their respective controls. The control efficacies and percent enhanced yields compared with their respective controls of the six bacterial endophytes were 29.6%～45.1% and 4.56%-10.59%, respectively. The strain JD204M had the highest capacities in controlling WSR and enhancing yields compared with other strains. The mechanisms on the suppression of WSR and the enhancement of yields on the wheat plants by the strains were futher tested.5. The induction of systemic resistance against WSR by strain JD204M was investigated. The results showed that the activities of phenylalanine ammonialyase (PAL), polyphenol oxidase (PPO), peroxidase (POD), catalase(CAT) and superoxide dismutase (SOD) were enhanced on the resistant cultivar (Xinyuan-958) and susceptible cultivar (Aikangdunchanwang) treated by the strain JD204M in the whole growth period compared to untreated control plants. The dynamic changes in these enzyme activities were significantly different between the two cultivars. The concentrations of malondialdehyde (MDA), O-2 and H2O2 were lower in the plants treated by strain JD204M compared to the controls, however, the concentrations of proline and flavonoid increased in the JD204-treated wheat plants. The results suggested the existence of systematic induction of resistance against WSR on the wheat plants treated by the endophytic bacterium.6. Effects of mutant JD204M on tiller number, plant height, root vigor and photosynthetic parameters on 14 different cultivars were investigated. The results showed that the tiller number of wheat seedlings (Zhenmai-9023) treated by the strain increased by 34.14%; the mean plant height and root vigor, chlorophyll content, and net photosynthesis rate (Pn), transpiration rate (7r) and stomatal resistance (Gs) in the wheat leaves increased by 9.21% and 13.33%,7.64% and 16.59%,10.74% and 20.32% on 12,10,11 cultivars, respectively, while the mean stomatal intercelluar CO2 concentration (Ci) decreased by 13.18% on 9 cultivars. The maximum quantum yield of PS II ((DPS II) on Aikangdunchanwang increased by 24.20%. The maximum photochemical capacity (Fv/Fm) and potential activity of PS Ⅱ (Fv/FO) increased by 19.45% and 10.36%, respectively. |
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