Engineering and Functional Expression of the Type III Secretion System in Xenorhabdus: Enhancing Insecticidal...

Area of Science:

• Microbiology
• Molecular Biology
• Insect Pathology

Background:

• Entomopathogenic nematode symbiotic bacteria (EPNB) are crucial for nematode-driven insect biocontrol.
• The type III secretion system (T3SS) is a key virulence factor in many bacterial pathogens.
• Understanding T3SS function in EPNB can unlock novel biocontrol strategies.

Purpose of the Study:

• To clone and functionally express the 32-kb T3SS gene cluster from *Photorhabdus luminescens* TT01.
• To evaluate the impact of heterologous T3SS expression on *Xenorhabdus stockiae* HN_xs01.
• To assess the potential of engineered EPNB for enhanced biocontrol efficacy.

Main Methods:

• Red/ET recombineering was used to clone the T3SS gene cluster.
• Functional expression of the T3SS was achieved in a T3SS-deficient *Xenorhabdus stockiae* strain.
• In vitro cell adhesion/invasion assays and *in vivo* insect models (*Helicoverpa armigera*) were employed.

Main Results:

• Heterologous T3SS expression significantly enhanced bacterial adhesion and invasion in CF-203 cells.
• Engineered bacteria caused severe intestinal damage in *H. armigera* by suppressing antimicrobial peptides.
• Improved bacterial colonization and a 3.7-fold decrease in LC50 were observed, indicating enhanced biocontrol.

Conclusions:

• The TT01-derived T3SS successfully delivered the novel effector XopA into host cells.
• Synthetic biology approaches can engineer EPNB for improved biocontrol capabilities.
• This study provides a mechanistic basis for T3SS-mediated biocontrol and future research.