Effects Of PH Value On Growth And Root Colonization Of Ralstonia Solanacearum

In recent years,soil acidification,which resulted in the serious occurrence of tobacco bacterial wilt,has gradually become obvious in China.Study on the relationship between pH value and occurrence of tobacco bacterial wilt can reveal the pathogenesis of tobacco bacterial wilt in acid soil and provide theoretical support for efficient agricultural control.In this paper,the pH range of R.solanacearum growth was determined,and the relationship between different pH value of soil and the incidence of tobacco bacterial wilt was tested.Secondly,the growth dynamics and gene copy number of R.solanacearum cultured in different pH of the NB medium and soil were determined by the method of qPCR.Besides these,the effect of pH on the pathogenesis-related factors,including Exopolysaccharide（EPS）and Lipopolysaccride（LPS）,film-forming ability and exercise intensity were measured.Finally,in different pH value conditions,the effect of EPS and LPS on adsorption onto the surface of tobacco roots or invasion into tobacco roots were determined by hydroponics,combined with scanning electron microscopy to observe the adsorption and colonization of R.solanacearum in tobacco roots.Main conclusions are as follows:1.The results showed that pH range of R.solanacearum growth was 5.2 to 8.0 cultured in NB medium under laboratory conditions.As the pot experiment results showing,soil pH is closely related to the occurrence of tobacco bacterial wilt.When pH within 5.5 to 6.5,the pH was positively related to the incidence of bacterial wilt,but pH within 6.5 to 8.0,the two relation was negative.When the soil pH was 6.0,6.5 and 7.0,the disease index of bacterial wilt was 42.72,56.60 and 42.20 respectively.As a result,the tobacco bacterial wilt occurred seriously in soils from weakly acidic to neutral（pH6.0⁷.0）.2.The results of qPCR indicated that the copy number of R.solanacearum gene cultured in weak acid NB medium is larger than those in neutral and alkaline conditions,that is,weak acid conditions are better for the growth of R.solanacearum;The dynamics of R.solanacearum was diverse in different pH tobacco-growing soils,the increasing rate of R.solanacearum in weakly acidic soil is faster than that in alkaline conditions.3.The results about effect of pH on pathogenesis-related factors showed that,the yield of EPS secreted by single and total colony of R.solanacearum were 3.07×10^-12 g and 52.28×10^-2 g cultured in acid NB medium（pH6.5）,respectively,which were higher than those which is cultured in neutral and alkaline conditions.The maximum total colony yield of LPS secreted by R.solanacearum was 9.03×10^-3 g at p H7.0,and the maximum single colony yield was 17.20×10^-15 g at pH7.5,the LPS yield in neutral and alkaline conditions were higher than that of acidic medium.The ability of biofilm formation was the strongest when pH was 5.5,followed by pH8.0,and pH7.0 was the minimum.;The motility of R.solanacearum cultured in different pH medium from strong to weak was:pH6.5,pH7.0,and pH8.0.The activity and biofilm-forming ability of R.solanacearum under weak acidity were stronger than that of neutral and alkaline conditions,which promoted the colonization and pathogenicity of R.solanacearum.4.Alkalinity affects the adsorption and invasion of R.solanacearum in tobacco roots.Under the different p H values conditions（5.2、5.9、6.6 and 7.3）,amounts of R.solanacearum adsorbing on the surface of tobacco roots after 24 h inoculation,were 6.50×10⁸、7.53×10⁸、11.6×10⁸ and 10.8×10⁸cfu/g﹒fw respectively,and the amounts of R.solanacearum invading into the root after 48 h were5.11×10⁹、5.90×10⁹、8.53×10⁹ and 7.97×10⁹ cfu/g﹒fw respectively.When we observed by SEM,the amount of R.solanacearum adsorbed on the surface of tobacco roots and white bacterial cells adhered to transverse section were the largest in the condition of p H6.6,followed by alkaline condition（pH7.3）,and that of p H5.2 was the minimum.The numbers of bacteria cell adsorbing and invading in tobacco roots were promoted by EPS and inhibited by LPS,but promoting effect of EPS played a leading role than that of LPS.