| Fusarium head blight (FHB) is a wordwide disease caused by Fusarium spp., also was the major diseases in wheat production of China. The wheat can be affected in the whole growth stage, and lead to kinds of seedling blight, stem rot, basal rot and head blight, which the most serious was the head blight. While with the pathogens’ resistance continuously strengthen, and there are more rainfall at flowering stage and pesticide residue pollution, have brought the huge challenge to the chemical control to continue to play an effective role. With the development of ecological pathology and comprehensive prevention and control measures, people considered using beneficial microorganisms which could increase production, prevent diseases, and protect the environment to control FHB. The biological control method has aroused widespread attention, and the purpose of this research is to screen microbial resources which could effectively control FHB and to explore its role mechanisms.282 microorganisms were screened from the body and rhizosphere soil of some plants, particularly the wheat.53 strains with antagonistic activity were obtained by the test of plate inhibition experiment. The strain XM5 from jointing stage of wheat leaf was selected for the research of colonization and disease resistance throughout the growing period of wheat. Two antagonistic strains PH5 and LHJ2 were isolated from wheat pollen and stem base respectively. A Streptomyces strain NS-QCT was stored in the laboratory, and through the field experiment prove that it has certain inhibition to FHB. This study was carried out on these three bacteria with the XM5 for the FHB research.An endophytic bacterial strain XM5 with strong inhibition to Fusarium graminearum was isolated from leaves of wheat at jointing stage. It was identified as Bacillus atrophaeus by analysis of 16S-23S internal transcribed spacer (ITS) nucleotide sequences. To develop an antibiotic-marker, the strain was cultured in the medium with two antibiotics rifampicin and streptomycin. Its antibiotic resistance to both of 100 μg·ml-1 rifampicin and streptomycin was induced by gradually increasing the concentration of antibiotics in the medium. To develop a DNA marker, we designed a pair of specific PCR primers L6SF and L6SR from a specific fragment of ITS sequence of XM5. With the double labels of the dual antibiotic resistance and the specific PCR test, the amount of XM5 colonized in wheat was analyzed in the indoor pot-culturing seedling and outdoor plant in different growth stages. The research found that XM5 could colonize in the wheat plant at different growth stages for a long time, its quantity could maintained at about 106 cfu·g-1 in root. The number of colonized XM5 in leaf showed downtrend along with growth time, although their was 2.63×106 cfu·g-1 XM5 in leaves at three-leaf stage, the number decreased to 104 cfu-g"1 at other stages. Comparing with root and leaf, less XM5 colonized in the stem and its number also decreased with time from 5.23×105 cfu·g-1 at three-leaf stage to 6×103 cfu·g-1 at jointing stage. Different applied methods of XM5 showed different results. We used the methods of seed soaking and root pouring with the bacterial suspension, and then investigated the colonization of XM5 in seedling and anti-pathogen effects on seedling blight. However, the result showed that only root pouring seedlings displayed disease resistance. The number of colonization in seedling showed a decreasing trend over time, which the number in 21 d decreased an order of magnitude than in 7 d. At different investigative times all colonized XM5 numbers presented as root>stem>leaf, for example, colonized XM5 in root was 3.10×106 cfu·g-1 in 7 d, it was 5.58 times more than in stem and 29.2 times than in leaf. The disease suppression index of the wheat seedlings showed the relative biological control efficiency was 45.65% in 14 d after root pouring. To research the biological control of XM5 to FHB, the bacterial suspension and culture broth were sprayed on the heading wheat, the number of endophytic XM5 in wheatear and the resistance efficiency against FHB were surveyed. The result showed that the number of XM5 on the surface of wheatear exhibited a down trend in both treatments, only less than 5% XM5 survived 10 d later. The endophytic number of colonized XM5 decreased greatly after 3 d for the spraying culture broth. It reduced to 5×102 cfu·head-1 when the time extended to 14 d. However, for the spraying bacterial suspension, the amount of colonized XM5 in the wheatears continuously increased from 5.2×103 to 4.8×104 cfu·head-1, and the biological control efficiency to FHB reached 68.3% in 7 d. These data showed that application of anti-pathogen bacteria XM5 to wheat could colonize in wheat plant. A suitable application method of antagonistic microorganisms on wheat heads could effectively control FHB. With the control effect, the strain exhibited best, XM5, NS-QCT took the second place, LHJ2 worst. From the colonization numbers, the reason may be the rapidly decline of the numbers of bacteria in the surface, but the specific reason relied on the further research and analysis. |
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