Role of Secondary Metabolites in Establishment of the Mutualistic Partnership between Xenorhabdus nematophila and the Entomopathogenic Nematode Steinernema carpocapsae

Abstract

bXenorhabdus nematophila engages in a mutualistic partnership with the nematode Steinernema carpocapsae, which invades insects, migrates through the gut, and penetrates into the hemocoel (body cavity). We showed previously that during invasion of Manduca sexta, the gut microbe Staphylococcus saprophyticus appeared transiently in the hemocoel, while Enterococcus faecalis proliferated as X. nematophila became dominant. X. nematophila produces diverse secondary metabolites, including the major water-soluble antimicrobial xenocoumacin. Here, we study the role of X. nematophila antimicrobials in interspecies competition under biologically relevant conditions using strains lacking either xenocoumacin (⌬xcnKL strain), xenocoumacin and the newly discovered antibiotic F (⌬xcnKL:F strain), or all ngrA-derived secondary metabolites (ngrA strain). Competition experiments were performed in Grace's insect medium, which is based on lepidopteran hemolymph. S. saprophyticus was eliminated when inoculated into growing cultures of either the ⌬xcnKL strain or ⌬xcnKL:F strain but grew in the presence of the ngrA strain, indicating that ngrA-derived antimicrobials, excluding xenocoumacin or antibiotic F, were required to eliminate the competitor. In contrast, S. saprophyticus was eliminated when coinjected into M. sexta with either the ⌬xcnKL or ngrA strain, indicating that ngrA-derived antimicrobials were not required to eliminate the competitor in vivo. E. faecalis growth was facilitated when coinjected with either of the mutant strains. Furthermore, nematode reproduction in M. sexta naturally infected with infective juveniles colonized with the ngrA strain was markedly reduced relative to the level of reproduction when infective juveniles were colonized with the wild-type strain. These findings provide new insights into interspecies competition in a host environment and suggest that ngrA-derived compounds serve as signals for in vivo nematode reproduction. Microorganisms employ various strategies to compete for space, nutrients, and other resources (1). In exploitative competition, limiting nutrients are quickly utilized by specific microorganisms without direct interaction between competitors. Interference competition, on the other hand, makes use of direct, antagonistic interactions. We study a tripartite model system in which the pathogenic bacterium, Xenorhabdus nematophila, establishes a mutualistic partnership with an entomopathogenic nematode, Steinernema carpocapsae, that vectors the bacterium into susceptible insect hosts. This tractable model system provides an excellent opportunity to identify biologically relevant competitors and study the role of X. nematophila antimicrobials in interspecies competition in a host environment.Xenorhabdus nematophila exhibits two distinct phases in its life cycle (2-5). It engages in a species-specific mutualistic relationship with S. carpocapsae, where it resides in a specialized region of the anterior intestine of the infective juvenile (IJ) stage of the nematode (6). The IJs invade s...

Entomopathogenic nematode-associated microbiota: from monoxenic paradigm to pathobiome

“…It is also generally asserted that X. nematophila outcompetes other bacterial strains or species during its lifecycle, particularly during the infestation phase of insect cadavers [20,45]. We therefore monitored the in vitro antibiosis of the X. nematophila XnSc_F1 strain against the cultivable bacterial members of the S. carpocapsae microbiota.…”

Symbiosis-inspired approaches to antibiotic discovery

“…31 Forst and co-workers have elucidated the role of xenocoumacins as well as other bacterially-derived NPs as signals important to in vivo nematode reproduction. 31 This is beyond the dual (and symbiotically advantageous) antibacterial and antifungal activities of Xcn1 and the more narrowly defined antifungal activity of Xcn2. 32 Enzymatic generation of Xcn2 from Xcn1 represents a putative mechanism by which X. nematophila avoids “self-toxicity” by limiting Xcn1 levels.…”

“…32 Notably, the X. nematophila genome houses six other NRPS-containing gene clusters and two large stand-alone NRPS genes; these BGCs drive the production of a wide array of NPs whose activities warrant continued study. 31 …”

Ready or Not: Microbial Adaptive Responses in Dynamic Symbiosis Environments

“…Finally, molecules produced by Xenorhabdus bacteria may themselves be preadaptive signals. Xenorhabdus secondary metabolites are produced during the reproductive phase of the life cycle and can antagonize invading microbial species to protect the insect cadaver (50,57). Changes in the local concentrations of these metabolites could be indicative of conditions that warrant IJ development and emergence.…”