Discovery of a microbially produced small molecule in a host-specific organ

The lifelong relationship between the Hawaiian bobtail squid, , and its microbial symbiont, , represents a simplified model system for studying microbiome establishment and maintenance. The bacteria colonize a dedicated symbiotic light organ in the squid, from which bacterial luminescence camouflages the hosts in a process termed counterillumination. The squid hosts hatch without their symbionts, which must be acquired from the ocean amid a diversity of non-beneficial bacteria, so precise molecular communication is required for initiation of the specific relationship. It is therefore likely that there may be specialized metabolites used in the light organ microenvironment to modulate these processes. To identify small molecules that may influence the establishment of this symbiosis, we used imaging mass spectrometry to analyze metabolite production in with altered biofilm production, which correlates directly to colonization capability in its host. ‘Biofilm-Up’ and ‘Biofilm-Down’ mutants were compared to a wild-type strain, and masses that were more abundantly produced by the biofilm-up mutant were detected. Using a combination of structure elucidation and synthetic chemistry, one such signal was determined to be a diketopiperazine, cyclo(-histidyl--proline). This diketopiperazine modulated luminescence in and, using label-free imaging mass spectrometry, was directly detected in the light organ of the colonized host. This work highlights the continued need for untargeted discovery efforts in host-microbe interactions and showcases the benefits of the squid- system for identification and characterization of small molecules that modulate microbiome behaviors.