Screening And Development Of Biological Agent Of Antagonistic Bacterium Against Colletotrichum Fructicola

Santalum album Linn.is one of the most expensive rare tree species in the world,it is useful and has huge economic value,cultivated in many southern regions of china.Anthrax is one of the main disease to affect the growth of sandalwood seedlings and sapling.At present chemical prevention is the main control method on sandalwood anthrax,there is no Antagonistic microorganisms control reports.In this paper,a antagonistic bacteria strain with highly antagonistic is screened from rhizosphere soil of Santalum album L.Identified and optimized fermentation process.The preliminary study on its wettable powder and field efficacy.(1)The screening and identification of antagonistic bacteria against Colletotrichum fructicola.Rhizosphere soil of superior individual plants of Santalum album L.was collected from Chengmai farm,71 bacteria strains were obtained by using dilution-plate method.From flat confrontation method screening and fermentation liquid compound sieve,one bacteria strain TXJ2-6 was found with highly antagonistic activity when Colletotrichum fructicola as the indicator bacteria.Results of Antibacterial spectrum tests showed that TXJ2-6 had obvious effect against Colletotrichum gloeosporioides,Agaricodochium camellia,FuSarium proliferatum,Pestalotiopsis microspora and Colletotrichum gloeosporioides.Finally,TXJ2-6 was identified as Bacillus methylotrophicus by its morphological,physiological and biochemical characteristics as well as 16s rDNA sequence and phylogenetic analysis.(2)The optimization of fermentation medium formula for Bacillus methylotrophicus TXJ2-6.We obtained a suitable fermentation medium through a singer-factor test:the carbon source is sucrose,the nitrogen source is yeast extract,inorganic salt is NaCl.targeted to OD600 of TXJ2-6 as response value,determine the point 0 level by steepest climbing test,These parameters further optimized using the Box-Behnken design experiment and response surface method(RSM),Finally,the optimal medium is sucrose 6.2g/L,yeast extract 13.0g/L,NaCl 2.0g/L.The reliability of the models was validated by validation experimentation.The optimization of fermentation conditions for Bacillus methylotrophicus TXJ2-6.The results of single-factor test and PB experiment showed that the initial pH,culture temperature and culture time were the key factors to influence the bacterial growth and inhibitory rate of TXJ2-6.Then devised the Box-Behnken experiment of 3 factors in 3 levers using the software Minitap 16.analyze the test data by response surface method(RSM),Finally,the optimal fermentation conditions is initial pH8.2,culture temperature 34.3 ?C and culture time 37h.The reliability of the models was validated by validation experimentation.(3)The development of TXJ2-6 WP.With the biocompatibility and antibacterial activity,the wetting time,suspension rate and stability as performance index,the optimal formulation of WP were confirmed as follows:original medicines70%,carrier of kaolin 10%,dispersing agent lignin sulfonate6%,wetting agent SDBS4%,stabilizer CMC-Na2%,UV protection agents ascorbic acid 0.1%,and make up to 100%with kaolin.All its indexes come up to the national and professional standards.The WP of TXJ2-6 can be stored at room temperature 25? and 4?,and can be stored for at least 4 months.Field experiments showed that the TXJ2-6 wetter powder had good effect on sandalwood anthracnose,and can be used as biological agents for control plant disease in the filed.