Comparative pathogenicity of entomopathogenic nematodes symbiotic bacteria- distinct intra- and inter-species variation.

This study aimed to achieve two main objectives: first, to determine whether the virulence factors of symbiotic bacteria of entomopathogenic nematodes (EPNs) against insect hosts are cell-associated or secreted, and to shed light on the underlying mechanisms of pathogenicity; and second, to identify and evaluate the standalone pathogenicity of symbiotic bacteria associated with entomopathogenic nematodes against Tenebrio molitor. Three bacterial species, Xenorhabdus nematophila (A41, SC, A18 and SF), Photorhabdus kayaii, and P. thracensis, were isolated and characterized via phylogenetic analysis of 16S-rRNA and gyrB genes. Bioassays involved injecting T. molitor larvae with bacterial suspensions, cell-free supernatants, immersing them in bacterial suspensions, and incorporating various bacterial treatments into their diet. Lethal concentrations (LC50) and lethal times (LT50) were determined based on observed mortality trends within specified periods. No mortality was recorded in T. molitor larvae when they were immersed in bacterial suspensions or fed diets incorporating various bacterial treatments. Among injection-based trials, X. nematophila strains A41, SC, and SF demonstrated remarkable virulence through low LC50 values in bacterial suspension tests, whereas X. nematophila A18 stood out in intracellular suspension trials. Interestingly, P. thracensis triggered mortality exclusively during extracellular suspension testing. However, no discernible effects emerged when employing non-invasive techniques such as soaking or spiking the insect's diet. Overall, our investigation highlights significant intra-species/inter-species variation in the pathogenicity profiles of nematode-associated bacteria towards T. molitor across diverse application modes, emphasizing the importance of tailored implementation approaches in utilizing microbials as effective biocontrol agents. The variation in susceptibility of host to cell suspension, cellular extract, and cell-free suspension of the studied bacteria indicates that the existence of living bacteria or intercellular secretions may be essential for certain isolates to establish a successful infection. In order to optimize the use of symbiotic bacteria as a biocontrol agent, it is important for future research to focus on identifying genes that encode for intercellularly secreted proteins and exploring alternate delivery methods. This will help in realizing the full potential of these beneficial bacteria in IPM programs.