Background: Photorhabdus are motile members of the family Enterobactericeae that are pathogenic to insect larvae whilst also maintaining a mutualistic interaction with entomophagous nematodes of the family Heterorhabditiae. The interactions between Photorhabdus and its hosts are thought to be an obligate part of the bacteria's life-cycle in the environment. Motility often plays a key role in mediating bacteria-host interactions and, in this study, we were interested in characterising the role of motility in the Photorhabdus-nematode-insect tripartite association. Results: We constructed deletion mutants of flgG (blocking flagella production) and motAB (blocking flagella rotation) in P. luminescens TT01. Using these mutants we show that both the ΔflgG and ΔmotAB mutants are equally as good as the wild-type (WT) bacteria in killing insects and supporting nematode growth and development suggesting that flagella production and motility are not required for pathogenicity or mutualism. However we show that the production of flagella is associated with a significant metabolic cost during growth on agar plates suggesting that, although not required for pathogenicity or mutualism, there must be a strong selective pressure to retain flagella production (and motility) during the interactions between Photorhabdus and its different hosts. To this end we show that both the ΔflgG and ΔmotAB mutants are out-competed by WT Photorhabdus during prolonged incubation in the insect revealing that motile bacteria do have a fitness advantage during colonisation of the insect larva. Conclusion: This is the first report of a role for motility in Photorhabdus and we show that, although not required for either pathogenicity or mutualism, motility does contribute to the competitive fitness of Photorhabdus during infection of the insect (and, to a lesser extent, the nematode). This adaptive function is similar to the role ascribed to motility in mammalian pathogens such as uropathogenic Escherichia coli (UPEC). Therefore, in addition to describing a role for motility in Photorhabdus, this study reinforces the relevance and utility of this bacterium as a model for studying bacteria-host interactions.