Identification Of The Antagnistic Bacterium JY1-5 And Biological Control Mechanism Of This Strain Against Botrytis Cinerea

Tomato grey mould is one of the important diseases in tomato greenhouse cultivation,which has caused great threat to the production of tomato.Antagonistic bacterium JY1-5 isolated from the rhizosphere of radish in Jiangyin city,Jiangsu province could inhibit mycelia growth of Botrytis cinerea and many other plant pathogenic fungi.Biocontrol potential of this strain was evaluated against the tomato grey mould disease in a greenhouse experiment.The classification status of JY1-5,the cultivation conditions for the production of antifungal substances and its identification were performed in this paper.We conducted the control of tomato gray mold disease in the agricultural sightseeing garden of Jiangwang in 2016 and 2017.The results showed that JY1-5 displayed more than 32%control effect on tomato gray mould which was better than that of procymidone.JY1-5 was tentatively identified as Paenibacillus polymyxa based on 16S rRNA sequence similarity,physiological and biochemical characteristics.JY1-5 grew slowly in TSB and LB media,and it's OD600nm were 0.30 and 0.29,respectively within 72 hours.The OD600nm were 1.59,1.39,1.37 and 1.07 within 72 hours when grown in Glucose-Starch-CaCO3(GSC),Landy,Katznelson and Lochhead(KL)and PDB media.Maximum growth was obtained in the NB culture medium,and the OD600nm could reach 1.66 within 72 hours.Antifungal activity of JY1-5 cell extract was observed in NB,PDB,KL and GSC media,and the inhibitory rates were 84.7%,85.2%,80.5%and 76.1%,respectively.The antifungal activity of culture supernatant filtrate were observed in KL,Landy and PDB media,and the inhibitory rates were 85.9%,81.9%and 88.6%,respectively.The culture supernatant filtrate of other media showed weaker antifungal activity.Stronger antifungal activity was observed in extracts from both culture supernatant filtrate and bacterium cells,when strain JY1-5 was cultured in KL and PDB media.Wheras the antagistic activity from cell extract was better than that from the culture supernatant,The effect of incubation time on the growth and antifungal substance production in KL medium were determined.Growth and antifungal activity of JY1-5 was found to be optimal at 36 h and decreased gradually in the following 12 h.The methanol extract of JY1-5 cells has broad antifungal spectrum including Fusarium graminearum,Magnaporthe grisea,Rhizoctorzia solani,Pythium aphanidermatum,Fusarium oxysporum,Gaeumannomyces gramini,Monilinia fructicola,Botrytis cinerea,Sclerotinia sclerotiorum,Erwinia carotorora,Xanthomonas campestris and Streptomyces scabies.The inhibition rate of cell methanol extract was over 80%in tomato fruit assay.The methanol extracts of JY1-5 cells were detected by MALDI-TOF/TOF mass spectrometry(MALDI-TOF/TOF-MS),which showed that molecular masses of m/z = 869.50,883.54,897.54,901.54,915.55,929.58,947.53,965.53,979.55.The antifungal compounds were members of the fusaricidin group of cyclic depsipeptides.The fragmentation patterns for 883.54 and 897.54 were consistent with the molecular structures of fusaricidin A and fusaricidin B according to the literature.Besides fusaricidin,the strain JY1-5 was able to produce extracellular degradation enzymes,including cellulase,chitinase,protease and amylase,but was not able to produce HCN and siderophore.It is not detected that JY1-5 has the function of phosphate solubilisation and nitrogen fixation.The above results provide an important reference value for the application of biocontrol strain JY1-5 against the tomato grey mould disease.