Screening,Identification Of Endophytic Bacteria From Citrus And Their Control Effect On Citrus Canker

[Research Purpose] Citrus is an important cash crop and its diseases are occuring seriously.Citrus canker caused by Xanthomonas axonopodis pv.citri,a quarantine disease named citrus canker,has brought serious economic loss to the citrus industry.At present,the disease is controlled by using the chemicals mainly that impeded the sustainable development of the citrus industry.Morever,the several chemicals applying are limited and absent in the research and exploit of new chemicals.It is kown that the long and frequent application of the same chemicals is able to lead the production of the pathogenic bacteria resistance to bactericides and the residual of the pesticides causing the pollution of environment and the threats of the human health.[Method](1)The endophytes were isolated from the tender leaves and fruits of Newhall navel orange summer shoots picked in different places in GanNan garden.(2)The endophytes against Xanthomonas axonopodis pv.citri were checked and screened by the methods of bioassay in plate primarily,and further the endophytes form citrus controlling the citrus canker were screened by the methods of inoculating the citrus leaves.(3)The endophytes screened were identified by morphological characteristics,cultural characteristics,physiological and biochemical characteristics,molecular biology method including 16 S rDNA gene sequence analysis and phylogenetic tree construction in order to clarify the classification status of strains.(4)By the use of the single factor experiment,the fermentation mediums were screened and the fermentation conditions including pH,fermentation time,inoculum were optimized,for endophytic bacterial strains which are able to control the citrus canker.(5)The effect of these strains identified on controlling the citrus canker were detected after inoculating the bacteria to the seedlings and adult trees of navel orange.[Result](1)The density of isolated colonies from one gram fresh weight of Gannan navel orange Newhall is 7.13×102colony form unit,and approximate 18 kinds of bacteria cultured are obtained.(2)3 kinds of bacteria including stain GN233?GN222?GN223 which are able to inhibit the growth of Xanthomonas axonopodis pv.citri and control the occur of citrus canker obviously are screened.(3)Strains GN222 and GN233 were identified as Bacillus subtilis and GN223 belonged to Enterobacter sp.by morphological characteristics,cultural characteristics,physiological and biochemical characteristics,molecular biology method including 16 S rDNA gene sequence analysis and phylogenetic tree construction.(4)Strain GN222 presented the best activity on Xanthomonas Campestris pv.citri when 1 percent of GN222 was inoculated in 50 ml of 250 ml TB fermentation nutrient solution with pH8 for60 hours,and strain GN223 presented the best activity on Xanthomonas Campestris pv.citri when 1 percent of GN223 was inoculated in 50 ml of 250 ml YPD fermentation nutrient solution with pH8 for 60 hours,and strain GN233 presented the best activity onXanthomonas Campestris pv.citri when 5 percent of GN233 was inoculated in 100 ml of250ml YPD fermentation nutrient solution with pH7 for 60 hours.(5)Some effects of 3strains against citrus canker were performed in the seedlings and adult trees of the navel orange,and strain GN222 which was superior to Streptomycin is best in all strains but GN223 is worse relatively.[Conclusion] 3 strains sreened are able to control the citrus canker and the effect of Bacillus subtilis GN222 with more the worth of application and research is the best.