Preliminary Stduy On Pathogen Identification And Control Of Stipe Rot Disease In Coprinus Comatus

Coprinus comatus is one of the important cultivated edible fungi in China.I ts unique taste,rich nutrition and medicinal value make it more and more pop ular with consumers.Coprinus comatus growth cycle is short,high bioconversi on rate,easy to cultivate.But the cultivation of pests and diseases in the proces s of serious,often causing serious economic losses.In particular bacterial disea ses occur quickly,spread quickly and dissemination of a wide range of prevent ion and treatment difficult.In this paper,we investigated the stipe rot disease of Coprinus comatus,and verified the pathogenicity of pathogens according to the Koch's law.The pathogen was identified,isolate pathogens and correspondi ng clinical standard strains were tested for animal pathogenicity.The soil sampl es were collected at different sites to analyze the source of the pathogen,scree ning plant extracts and biocontrol bacteria to carry out the prevention and cont rol program.To ensure the healthy development of the industry,further control of bacterial diseases of Coprinus comatus,laid the foundation for food safety work.JTG-B1,JTG-B2 and JTG-B2 were obtained by isolation and purification in the breeding ground of Coprinus comatus in Pingyin County,Shandong Province in July2015.According to the Koch's law,the experiment was conducted to verify the pathogenicity of the three isolates.The inoculation method of the three isolates was as follows,according to whether the wear and inoculation is divided into four groups,each group of base for the 25 healthy Coprinus comatus fruit and repeated 4 times,The symptoms and number of fruiting bodies were observed at 16 hour,36 hour and52 hour.The results show that JTG-B1 can cause half of the fruiting body in 16 hours,and there are obvious lesions and bacterial pus appear late in the latter part of the serious start of the fruiting body until death.Therefore,the pathogenicity of JTG-B1 was significantly stronger than JTG-A and JTG-B2,and the number of pathogens was more than other two strains.The morphological observation,physiological and biochemical reactions and 16 s rDNA of JTG-B1 were identified as Achromobacter xylosoxidans.JTG-B1 and TJ-ade were used to confirm whether the JTG-B1 and TJ-ade had animal pathogenicity.The mice were treated with JTG-B1 and TJ-ade from the Tongji Hospital affiliated to Huazhong University of Science and Technology.The results showed that 80%JTG-B1 suspension and 20% saline mixture intraperitoneal injection of mice after 21 d eating rate was 33%,TJ-ade the same treatment was 29%.Mice in the treatment group were no phenomenon of death,the mice was not active,the eyes closed with embrace phenomenon,body hair off.The optimum pH and temperature of JTG-B1 and TJ-ade strains were measured.JTG-B1 and TJ-ade could grow at 16?to 34?,and the optimum growth temperature at 28?.Between pH 4 and 11 both can grow,at 7 is the most appropriate growth pH.Amplification of JTG-B1 and TJ-ade's seven housekeeping genes,nusA,rpoB,eno,gltB,lepA,nuoL,nrdA,were sequenced and the MLST database was compared withthe allele number.After PCR reaction,agarose gel electrophoresis showed that the size of the stellate gene was 300-400 bp,and JTG-B1 and TJ-ade were ST14 and ST29,respectively.TJ-ade in the 16 h when the injury group can also be the same with JTG-B1 half of the physical disease,the same symptoms for JTG-B1.CIT(citrate test),VP(pyruvate test),GLU(glucose fermentation test),SAC(sucrose fermentation test),AMY(amygdalin fermentation test),ARA(arabinose test)as a positive result.However,the color reaction of JTG-B1 was similar to that of TJ-ade,and the metabolic response rate and the expression of related enzymes were higher than those of TJ-ade.The pathogenicity experiment was combined to analyze the two species Differences between the situation.The soil samples were extracted from the seven locations around the Coprinus comatus cultivation base in Pingyin County,Shandong Province,and the total DNA was extracted and the blaOXA-114 gene of the target pathogen JTG-B1 was amplified.The PCR products of the ground outside the cave were analyzed by electrophoresis.There is a band of about 500 bp in size,to prove that the pathogen may be outside the cultivation site of Coprinus comatus through human activities,not completely sterilized cultivation of mosquitoes or mosquitoes into the cultivation environment.Preparation of garlic,sycamore leaves,ginkgo biloba,busax,solanum,tea,cinnamon oil,pomegranate peel,pepper extract.The results showed that only cinnamon oil concentration of 100 mg / mL and 120 mg / mL,the pathogen can produce the size of 1.0 mm and 9.0 mm inhibition zone.bacteria fermentation of Pseudomonas fluorescens and Pseudomonas putida broth no effect on pathogens.The cinnamon oil with the concentration of 120 mg / mL had no effect on the mycelial growth of Coprinus comatus.The cinnamon oil could also play a role in the prevention and treatment of pathogens at 12 hour and 24 hour.