Identification And Biological Control Of The Pathogen Of Ginger Rhizome Rot

Ginger rhizome rot is a kind of soil borne disease with great harm in ginger production.It is mainly caused by Pythium spp.and Fusarium spp.,and sometimes caused by bacteria.It reduces 50%-80%yield of ginger in some areas.In most cases,this kind of disease occurs in the years when weather conditions are proper for the growth of the pathogens.At present,chemical control is the main measure to deal with this disease,but excessive use of chemical pesticides will seriously affect human health and cause environmental pollution,and may produce drug resistance of pathogens.It has become the consensus of researchers and agricultural pesticides users to find broad-spectrum,efficient and low-toxic biological control methods.In the summer of 2018,there was a continuous strong rainfall in Weifang,Shandong province,which caused the rainwater to submerge most of the ginger planting plots,and the temperature turned higher after the rain.After the rainfall,the ginger rhizome rot broke out in a large number of areas,most of the ginger planting areas was infected,and the farmers lost a lot.Therefore,samples of diseased ginger were collected in September 2018,the casual agent of ginger rhizome rot was isolated and identified.The antifungal effect of antimicrobial lipopeptides derived from Bacillus spp.on the pathogen was studied,and the control effect of the lipopeptide which exhibited the highest inhibition rate against the pathogen was measured in vitro;in addition,an antagonistic Bacillus strain was screened by the method of dual culture,and the antagonistic effect on the pathogen of ginger rhizome rot was determined by co-culture in liquid medium;the sporulation medium and the parameters of culture conditions was optimized;finally,the control effect of the antagonistic Bacillus was measured in vitro.1.The pathogen of the rhizome rot of ginger was isolated from diseased ginger by tissue isolation method.The symptoms of ginger which was inocululated by the pathogen was same as infected ginger,the fungi was isolated from inoculated ginger again.So the isolated fungi met the Koch's rule.The pathogen was identified as Pythium myriotylum by morphological identification and molecular biological identification.2.All of the five antimicrobial lipopeptides have antifungal effect on Pythium myriotylum.Brevibacillin derived from Brevibacilluslaterosporus fmb 70 exhibited the largest inhibition rate against the Pythium myriotylum,and the EC50 of brevibacillin to Pythium myriotylum was determined as 13.081 ?g/mL;the growth and development of Pythium myriotylum was delayed after treated by brevibacillin;and it was observed that the peptide changed the membrane permeability of Pythium myriotylum;and it was found that the surface of Pythium myriotylum was wrinkled after treated by brevibacillin.In soil environment,the incidence of root rot in ginger slices decreased by 40%,56.7%,and the disease index decreased by 41 and 58.9 after treated by brevibacillin at concentrations of 26?g/mL and 52 ?g/mL,respectively.3.An Bacillus B3 exhibited antagonistic effect against Pythium myriotylum was obtained by screening,and was identified as Bacillus paralicheniformis by morphological observation,molecular biological identification and physiological biochemical identification.The antagonistic character against the pathogen was studied by co-culture method.The results showed that the weight of mycelium of Pythium myriotylum was only 9.2%of that of the control group when the initial concentration of Bacillus paralicheniformis B3 was 104 CFU/mL in PDB medium;when Pythium myriotylum occupied the nutritional advantage by pre-culture by 48 h in PDB before inoculation of Bacillus paralicheniformis B3,the growth of Pythium myriotylum was significantly inhibited by Bacillus paralicheniformis at the initial concentration of 105 CFU/mL.When the concentration of the antagonistic bacteria was 106 CFU/mLand 107 CFU/mL,the growth of Pythium myriotylum was inhibited by 88.5%and 93.1%.And it was observed by optical microscope that the Bacillus paralicheniformis B3 changed the surface morphology of Pythium myriotylum and the mycelium growth was curved,atrophic and deformed and uneven in thickness.4.The sporulation medium and the parameters of culture conditions was optimized,the optimized formula of sporulation medium was:wheat bran 10 g,yeast extract 14 g,calcium chloride 2.5 g,manganese sulfate 0.15 g,water 1 L;the best culture condition parameters:pH was 5.5,liquid loading was 30 ml/250ml,culture time was 72 h,the culture temperature was 40?.The number of spores of Bacillus paralicheniformis B3 increased from 4.6×106 CFU/mL to 9.55×109 CFU/mL by optimizing of culture medium and fermentation conditions.The control effect of the antagonistic bacteria in vitro was measured,the result showed that Bacillus paralicheniformis B3 could significantly reduce the incidence rate and infection index of ginger rhizome rot compared with control group:In soil environment,the incidence of root rot in ginger slices decreased by 43.3%,53.3%,and the disease index decreased by 67.7 and 71.7 after treated by Bacillus paralicheniformis B3 at the concentration of 107 CFU/mL and 108 CFU/mL,respectively.