[P007] Poster

12:10 PM - 1:00 PM JST(3:10 AM - 4:00 AM UTC)

[P04-467] Effects of Carbon Sources on Outer Membrane Vesicle-Related Particle Characteristics, and Insecticidal Activity of Photorhabdus luminescens

○Chienyan Hsieh¹ (1. National Kaohsiung Normal University (Taiwan))

Keywords:

Photorhabdus luminescens,outer membrane vesicles(OMV),Plutella xylostella,insecticidal activity,SDS-PAGE

This study investigated the effects of different carbon sources on extracellular high-molecular-weight proteins, outer membrane vesicle (OMV)-enriched particle characteristics, and insecticidal activity of Photorhabdus luminescens, and further examined the relationships among these factors. Three P. luminescens strains, 0813s, 2103, and ATCC, were cultivated in LB medium supplemented with 2% sorbitol, 2% sucrose, or 2% glycerol. Samples were collected after 72 h for comparative analyses. Extracellular proteins were analyzed by SDS-PAGE, with particular emphasis on the >100 kDa region as an indicator of putative high-molecular-weight insecticidal proteins. Nanoparticle tracking analysis (NTA) was used to characterize the concentration and size distribution of OMV-enriched particles, with special attention to the dominant 50–300 nm range. Insecticidal activity was evaluated using supernatant and whole-broth samples against third-instar larvae of Plutella xylostella, and 72 h mortality was used as the primary endpoint.The results demonstrated clear strain- and carbon source-dependent differences in extracellular protein profiles, OMV-enriched particle characteristics, and larvicidal activity. SDS-PAGE analysis showed that strain 0813s exhibited the highest enrichment of the >100 kDa protein fraction, with an overall trend of 0813s > ATCC > 2103. Among the carbon sources, glycerol produced the highest >100 kDa/total lane intensity ratio, whereas sucrose tended to increase the absolute abundance of the >100 kDa fraction under some conditions. NTA analysis showed that 0813s had the most stable OMV-enriched particle concentration across treatments, 2103 was the most responsive to carbon source changes, and sorbitol markedly enhanced particle release in 2103. In contrast, ATCC produced comparatively larger OMV-enriched particles overall. Bioassays showed that 0813s exhibited the strongest insecticidal activity, particularly under glycerol and sorbitol supplementation. At 25-fold dilution, both 0813s-sorbitol and 0813s-glycerol whole-broth samples achieved 100% mortality.Correlation analysis further revealed that the >100 kDa/total lane intensity ratio showed a moderate positive correlation with the 72 h mortality of supernatant samples, whereas the association between OMV-enriched particle concentration and insecticidal activity was weak. These findings indicate that enrichment of the high-molecular-weight protein fraction is a better predictor of soluble insecticidal activity than OMV-enriched particle abundance alone. However, because whole-broth samples frequently outperformed supernatants, the final insecticidal effect was likely not determined solely by soluble proteins, but also involved OMV-related or cell-associated factors. Overall, this study demonstrates that carbon source composition simultaneously affects extracellular protein production, OMV-enriched particle characteristics, and insecticidal activity in P. luminescens. Among the tested conditions, the combination of strain 0813s and glycerol showed the greatest potential, supporting the hypothesis that both high-molecular-weight proteins and OMV-related systems contribute to insecticidal function.