Establishment and application of efficient nematicidal screening model in Bacillus amyloliquefaciens

Abstract:

Abstract: ［Objective］ The mechanisms of nematophagous bacteria against nematodes remain unclear，limiting the use of biocontrol bacteria in the agriculture． Therefore，we constructed a rapid and efficient screening model to quickly identify new candidate genes involved in nematode infection.［Methods］ The wild-type Bacillus amyloliquefaciens FZB42 as well as more than 400 random mutants were inoculated into 5 mL liquid bioassay medium． After growth at 37℃ for 24 h，200 μL bacterial culture and 50－60 Level 4 age nematodes were added to 24-well plates，and then the survival rates of nematodes were determined at different time points．Through several rescreening，we selected the mutant strains whose nematicidal activities significantly decreased compared with the wild-type strain．Meanwhile，the conventional bioassay of solid plate was used as control.［Ｒesults］ Two mutants (F1 and F2) with obvious decreased nematicidal activities were selected from the random mutation library by liquid bioassay，consistent with the result of conventional solid plate bioassay. By comparing to 168 h-screening in each round of the solid plate bioassay，the method of liquid bioassay required only 24 h． The result indicated that the liquid bioassay greatly reduced the experimental time.［Conclusion］Our current study has successfully constructed a rapid and efficient method to bioassay the nematicidal activity，which could also lay the foundation for further cloning the candidate genes involved in the microbial infection against nematodes.