Analysis of the regulation and role of fimbriae of Xenorhabdus nematophila and release process from the intestinal receptacle of Steinernema carpocapsae

The aims of this study were to further understand the regulation of mrx fimbrial genes and to assess the function of Mrx fimbriae in the colonization of the nematode, and to elucidate the anatomical structure of the anterior intestine colonized by X. nematophila and detail how the bacteria move out of this structure into the intestine.Xenorhabdus nematophila is a symbiont of the entomopathogenic nematode Steinernema carpocapsae and is pathogenic towards diverse insects. It produces fimbriae composed of the major subunit MrxA when grown on Nutrient broth agar (NB-agar). We show that X. nematophila grown on Luria-Bertani broth agar (LB- agar) lack fimbriae and produce little MrxA. However, expression of mrx genes in cells grown on LB-agar was comparable to cells grown on NB-agar. Moreover, mrxA expression in a phenotypic variant cell that lacks fimbriae was similar to that of the wild type cell. These findings indicate that Mrx fimbriae production can be controlled at the post-transcriptional level. We further show that nematodes propagated on X. nematophila lacking fimbriae (LB-agar) were colonized to the same extent as nematodes grown on bacteria producing fimbriae (NB-agar). In addition, mrx mutant strains and phenotypic variant strains devoid of fimbriae colonized nematodes as well as wild type cells both in vitro (on bacterial lawns) and in vivo (in insects). Furthermore, mrx mutant strains did not show a competitive colonization defect in vitro . Thus, production of fimbriae was not required for nematode colonization per se. In contrast, co-injection of insect host with both the mrx mutant and wild type strains resulted in nematode progeny that were colonized almost exclusively with the wild type strain indicating that fimbriae confer a competitive advantage in vivo. These findings reveal novel aspects of both regulation and function of the Mrx fimbriae of X. nematophila.Previous studies suggested that X. nematophila colonizes a vesicle structure connected to the anterior intention of the infective juvenile (IJ) stage of Steinernema carpocapsae. Here we show that Xenorhabdus colonizes the distal part of an expanded region of the anterior intestine called the receptacle and not a separate vesicle structure. Exposure to insect hemolymph stimulated pumping of the basal bulb structure in the nematode and pulsatile movement of the bacteria in the receptacle. The pulsatile movement continues until the bacteria move posteriorly into the intestinal lumen. A non-motile strain was released efficiently, suggesting that bacterial motility is not required for movement out of the receptacle. Agents that induced nematode paralysis dramatically reduced bacterial release indicating that the pumping action of the basal bulb provides the force for bacterial release.