Entomopathogenic bacteria of the genus Xenorhabdus are known to be symbiotically associated with soil dwelling nematodes of the Steinernematidae family. These bacteria are transported by their nematode hosts into the hemocoel of the insect larvae, where they proliferate and produce insecticidal proteins, inhibitors of the insect immune system and antimicrobial molecules. In this study, we describe the discovery of a new family (PAX) of five antimicrobial compounds produced by fermentation of the Xenorhabdus nematophila F1 strain and purified by cation exchange chromatography and reversed phase chromatography. The chemical structure of PAX 3, a lysine-rich cyclolipopetide, was obtained from the analysis of homo and heteronuclear 2D NMR and confirmed by MS-MS experiments. The five members of the PAX family showed significant activity against plants and human fungal pathogens and moderate activity against few bacteria and yeast. No cytotoxicity was observed on CHO or insect cells.
Xenorhabdicin, the phage tail-like bacteriocins of Xenorhabdus nematophilus, and phage head particles, elements produced together after mitomycin induction in X. nematophilus lysogenic strain F1 cultures, were separated by DEAE chromatography, examined by transmission electron microscopy, and characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Electrophoresis of xenorhabdicin showed two major subunits of 43 and 20 kDa corresponding to the sheath and the inner core, respectively. At least five other minor subunits of 67, 54, 35, 28, and 16 kDa were also characterized. Electrophoresis of the phage head capsids showed a major 40-kDa subunit and two minor 50-and 34-kDa subunits. Bactericidal activity recorded against closely related bacterial species and spontaneously produced by X. nematophilus resides in the xenorhabdicin particles and is another antimicrobial barrier to save the symbiotic association.
Induction by mitomycin or high-temperature treatment resulted in the production of bacteriocins and phages in both phases of Xenorhabdus nematophilus A24, indicating lysogeny. Phage DNA purified from X. nematophilus A24 hybridized to several fragments of DraI-digested A24 chromosomal DNA, confirming that the phage genome was incorporated into the bacterial chromosome. Bacteriocins and phages were detected in cultures of most other Xenorhabdus spp. after mitomycin or high-temperature treatment. Xenorhabdus luminescens K80 was not lysed by these treatments, and no phages were seen associated with this strain. However, bacteriocins were detected in limited quantities in all Xenorhabdus cultures, including X. luminescens K80, without any induction. X. nematophilus A24 bacteriocins were antagonistic for other Xenorhabdus species but not for A24 or other strains of X. nematophilus.
Xenorhabdus spp. and Photorhabdus spp., entomopathogenic bacteria symbiotically associated with nematodes of the families Steinernematidae and Heterorhabditidae, respectively, were shown to produce different lipases when they were grown on suitable nutrient agar. Substrate specificity studies showed thatPhotorhabdus spp. exhibited a broad lipase activity, while most of the Xenorhabdus spp. secreted a specific lecithinase. Xenorhabdus spp. occur spontaneously in two variants, phase I and phase II. Only the phase I variants ofXenorhabdus nematophilus and Xenorhabdus bovienii strains produced lecithinase activity when the bacteria were grown on a solid lecithin medium (0.01% lecithin nutrient agar; 24 h of growth). Five enzymatic isomers responsible for this activity were separated from the supernatant of a X. nematophilus F1 culture in two chromatographic steps, cation-exchange chromatography and C18 reverse-phase chromatography. The substrate specificity of the X. nematophilus F1 lecithinase suggested that a phospholipase C preferentially active on phosphatidylcholine could be isolated. The entomotoxic properties of each isomer were tested by injection into the hemocoels of insect larvae. None of the isomers exhibited toxicity with the insects tested, Locusta migratoria, Galleria mellonella, Spodoptera littoralis, and Manduca sexta. The possible role of lecithinase as either a virulence factor or a symbiotic factor is discussed.
Summary— Xenorhabdus nematophilus FI strain and Photorhabdus luminescens NC19 strain produced bacteriocins after mitomycin C treatment and under natural conditions respectively. The ultrastructure of these two strains was described and compared to the ultrastructure of untreated or normal cells. After image processing of purified bacteriocins we found morphological homology in infected cells with protoplasmic rods in longitudinal section and hexagonal aggregates in transversal section. We concluded that these particular structures, so‐called ‘lattice structures’ and previously interpreted as ‘photosomes’, are in fact the early stages of in situ production of bacteriocins in these two bacterial genera. Natural occurrence of Photorhabdus spp bacteriocinogenesis was observed in other strains, while other lysogenic strains of Xenorhabdus spp are lysed after a mitomycin C treatment.
Analysis of the Photorhabdus luminescens genome sequence revealed that the pts region is related to the tail synthesis gene core of the P2 phage. The pts locus encodes a DNA invertase homologue. PCR-RFLP analysis showed the two potential tail fiber regions of the pts locus present DNA inversions. Electron microscopy revealed a phage tail-like particle, related to the R-type family and named R-photorhabdicin, in the culture supernatant of P. luminescens. Mass spectrometry analysis of two sub-units of R-photorhabdicin revealed that they are encoded by the pts locus. The role of this P2-related prophage remnant in the Photorhabdus genome is discussed.
Peptides U 0400 Identification of a New Antimicrobial Lysine-Rich Cyclolipopeptide Family from Xenorhabdus namatophila. -(GUALTIERI*, M.; AUMELAS, A.; THALER, J.-O.; J. Antibiot. 62 (2009) 6, 295-302; Fac. Pharm., CNRS, Univ. Montpellier I, F-34093 Montpellier, Fr.; Eng.) -H. Toeppel 38-187
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.