Identification and Nematicidal Characterization of Proteases Secreted by Endophytic Bacteria Bacillus cereus BCM2

Abstract

The endophytic bacterium Bacillus cereus BCM2 has shown great potential as a biocontrol organism against Meloidogyne incognita, which causes severe root-knot diseases in crops. In our previous study, the metabolite of BCM2 showed high nematicidal activity against the M. incognita second-stage juveniles. However, the mechanism employed by endophytic bacteria to infect and kill nematodes is still unclear. Here, we investigate both the endophytic bacterial extracellular proteins with nematicidal activity and their mechanism of killing nematodes. The first step was detecting the nematicidal activities of crude proteins. The results show that the nematode mortality rate reached 100% within 72 h, and the crude proteins damaged both the cuticle and eggshell, before finally destroying the targets. This suggests possible proteinaceous pathogeny in BCM2. Throughout the process, the fine-detail changes in the nematode cuticle and the intestinal structure were observed using scanning electron microscopy and transmission electron microscopy. These images show that BCM2 extracellular proteins did not damage the internal organization of the nematode but did severely damage its cuticle, which led to content leakage. From the crude proteins, chitosanase, alkaline serine protease, and neutral protease were purified and identified. The M. incognita–B. cereus BCM2 microenvironment simulation demonstrates that BCM2 adheres to the surface of nematodes and helps the metabolites that were produced by BCM2 to rapidly recognize and kill M. incognita. This relationship between plants, endophytic bacteria, and nematodes offers insight into the biological mechanisms that can be utilized for of nematode management.