Chemotaxis Response Of Ginseng Bacterial Soft Rot Pathogen To Three Terpenoid Saponins From Panax Ginseng And Its Pathogenic Mechanism

Ginseng(Panax ginseng C.A.Mey)belonging to Araliaceae is a kind of perennial medicinal plant,cultivated for the high medicinal value of its root.It has the functions of nourishing vital energy,Fumai solid-off,nourishing spleen,nourishing the lungs,nourishing blood,nourishing and stimulating the nerves.The genuine producing area of ginseng is located in northeast.Ginsenosides are the material basis of ginseng medical and pharmaceutical functions.In recent years,with the adjustment of agricultural industrial structure and the expanding of the cultivation area of farmland ginseng,ginseng bacterial soft rot(Erwinia carotovora subsp.carotovora(Jones)Bergey et al.),one of the important disease in the production of ginseng in China,has became the restriction on the healthy development of ginseng industry,which has greatly affected the output and quality of ginseng.So far,there is little systematic research between host and host-pathogen interaction mechanism at home and abroad.Based on the principle of plant pathology,this paper used physiological and biochemical techniques to explore the chemotactic response of ginseng bacterial soft rot to ginseng triterpenoid saponins and the pathogen's pathogenic mechanism,in order to scientifically control the bacterial rot of ginseng.1.Chemotaxis response of Bacterial soft rot to ginsenosidesChemotaxis is the first step in the pathogenes was of some pathogens,and is the initial manifestation of mutual recognition.The chemotaxis response of bacterial soft rot of ginseng to ginsenosides was investigated by capillary method and selected the optimal chemotaxis conditions.The optimal chemotaxis concentrations of bacterial soft rot of ginseng to total ginsenosides,panaxadiol glycosides and protopassin glycosides were concentration 5 mg/L,and the optimal chemotaxis concentration of Rd and Re was 0.5 mg/L.The optimal chemotaxis p H value of bacterial soft rot of ginseng to the panaxoglycol-type aglycone,protopanaxaglycoside-type aglycone and Re was 7 mg/L.The optimum chemotaxis p H value of the total ginsenosides and Rd was 8.The optimum chemotaxis temperature and time for these five kinds of chemo attractants were 30? and 60 min,respectively.Ginsenosides could induce chemotaxis in ginseng bacterial soft rot.The same method was used to analyze the chemotaxis response of ginseng bacterial soft rot to ginseng root exudates.The maximum chemotaxis value of the ginseng root exudates in the aqueous layer of ginseng at p H=7 and concentration of 0.0125 mg/L was obtained at 25 °C for 60 min.The chemotaxis rate was 2.2519,and the p H value was 7.At the concentration of 0.125 mg/L n-butanol and petroleum ether ginseng root exudates reached the maximum chemotaxis value at25 °C and 30 °C for 60 min and 75 min,respectively,and the chemotaxis rate was2.40,2.163.A series of proteins and chemotaxis pathways involved in the complete chemotaxis of genomic DNA at the genomic level of E.carotovora subsp.carotovora were obtained,and the chemotaxis domain of the bacteria was predicted.It could be concluded that the accumulation of ginseng root exudates could attract ginseng bacterial soft rot pathogens to accumulate around the roots of ginseng,which greatly increased the chance that the bacterium would successfully infect ginseng.2.Pathogenicity and microstructure analysis of ginseng roots infected by ginsengbacterial soft rot pathogen.By carring out a test about the pathogenicity of the collected bacterial strains of bacterial soft rot pathogens,we found the pathogenicity among the different strains was obviously different.The order of pathogenicity was: ginseng bacterial SN-1>SN-3> JG-2> JG-1> JN-1> SN-2.Microscopic observation showed that the bacterial soft rot pathogen invaded into the wound of ginseng root epidermis cells and showed radial infection.As the culture time prolonged,the soft rot pathogen gradually developed into the periderm,phloem,and formation layers,and the xylem successively expanded and proliferated.Finally,the root tissue showed typical symptoms of brown soft rot.Ginsenosides could delay and prevent the course of bacterial infection when ginseng bacterial soft rot pathogen infected,and ginseng saponin had the ability to kill individual bacteria.Since ginseng root pericarp was continuously protected by ginsenosides which could be used against ginseng bacterial disease outbreaks.3.Effects of ginsenosides on cell wall degrading enzymes produced by ginseng bacterial soft rot bacteriaGinsenosides aglycon could promote the enzymatic activity of cellulase,polygalacturonic acid trans-elimination enzyme,and pectin methyl trans-elimination enzyme,and inhibited the enzymatic activity of pectin methylgalacturonase.Within 24 hours after infection,ginsenosides aglycon can promote polygalacturonase and cellulase in pathogens,and ginsenosides can promote cellulase at 36 hours after infection.During the process of infection,ginsenoside and ginsenosides aglycon stimulated the activity of polygalacturonate trans-elimination enzyme and pectin methyl trans-eliminase.Ginsenosides and ginsenosides aglycon had different effects on cell wall degrading enzymes produced by ginseng bacterial soft rot pathogens.4.Ginseng' defense response induced by ginsenosidesGinseng bacterial soft rot pathogen invaded ginseng,ginsenoside reduced the O2-content in the first 12 hours,accelerated the ·OH clearance rate in the first 24 hours,postponed the ginseng from soft rot,and panaxsapogenol increased O2-content,accelerated of the ·OH clearance rate in the first 24 h.SOD enzyme activity showed that in the early stage of pathogen infection,ginsenosides could prevent bacterial infection;the latter role was not obvious.The panaxsapogenol not only did not stop the infection of the bacteria during the first 18 hours,but also aggravated the infection of the bacteria.Ginsenoside could increase the activity of CAT enzyme and prevent the infection of pathogens,while the effect of panaxsapogenol on CAT enzyme activity was not significant.Both ginsenosides and panaxsapogenol could increase POD activity in pathogen infection,and panaxsapogenol was stronger than saponin.At the late stage of pathogen infection,ginsenosides could significantly reduce MDA content,while panaxsapogenol increased MDA content.Ginsenoside could increase soluble protein content,while panaxsapogenol had no similar effect.At the same time,with the advancement of the invasion time of the bacteria,the content of free Pro in the plant decreased at the later stage of the ginsenoside course,which weakened the stress on the external stress.Panaxsapogenol exacerbatedthe adverse effects of pathogen infection.Ginsenoside could rapidly increase the GSH content in plants,but panaxsapogenol were not so sensitive.The ginsenosides and ginsenosides panaxsapogenol had significant differences in the activities of different antioxidant enzymes,and the differences were also significant at different stages of the disease course.Ginsenosides could reduce membrane lipid peroxidation and delay and attenuate invasion of Ginseng bacteria.Panaxsapogenol could exacerbate ginseng membrane lipid peroxidation and aggravate the infection of pathogens.5.The response metabolomic analysis of bacterial soft rot of ginseng to ginseng triterpenoid saponinThe continuously stimulating of ginsenosides and ginsenosides aglycones during the infection process of ginseng bacterial soft rot pathogens was monitored by metabolomics method.In the infection of ginseng bacterial soft rot,the species of substances in ginseng tissue increased by using UHPLC-QTOF-MS for detection of positive and negative ion patterns.Compared to the sterile water treatment group(s2),1 organic acid,1 ketone,1 glycoside,4 lipids,2 nucleotides,and 12 amino acids were increased in the ginsenoside treatment group(s3),while 5 nucleosides,6organic acids,1 lipids and 1 alcohols,carbohydrates 1,alkaloids 1,nucleotides 4,amino acids 2were decrease.There were 3 organic acids,1 polysaccharide,and 3 amino acids increased in the ginsenoside aglycones treatment group(s4)compared to s2 and 1 species of carbohydrates,1amino acid,and 1 organic acid were decrease.For the qualitative and accurate comparison of the content of different substances,the ginsenosides were screened for compounds with significant differences.