A lasting symbiosis: how Vibrio fischeri finds a squid partner and persists within its natural host

Abstract

As our understanding of the human microbiome progresses, so does the need for natural experimental animal models that promote a mechanistic understanding of beneficial microorganism-host interactions. Years of research into the exclusive symbiosis between the Hawaiian bobtail squid, Euprymna scolopes, and the bioluminescent bacterium Vibrio fischeri have permitted a detailed understanding of those bacterial genes underlying signal exchange and rhythmic activities that result in a persistent, beneficial association, as well as glimpses into the evolution of symbiotic competence. Migrating from the ambient seawater to regions deep inside the light-emitting organ of the squid, V. fischeri experiences, recognizes and adjusts to the changing environmental conditions. Here, we review key advances over the past 15 years that are deepening our understanding of these events.

Fig. 1.. The juvenile E. scolopes light organ.

a. The nascent light organ (circle) is located in the mantle cavity of a newly hatched squid. b. Two pairs of ciliated appendages emerge from the outer surface of the light organ, interacting with seawater drawn into the mantle cavity during respiration. c. Three pores are found at the base of each pair of appendages. d. Soon after hatching the appendages begin to produce mucus that covers the ciliated fields. e. Mucus production and the flow fields created by the cilia capture V. fischeri cells (here, labelled with GFP) present in the seawater, and direct them to a zone directly above the pores where they form aggregates. f. After a few hours, the aggregates chemotax to the three pores (square), and migrate into one of three interior ducts, each leading to an antechamber (a), through a bottleneck, and into a crypt. Once in the crypts, the V. fischeri cells proliferate and autoinduce luminescence. This interior pathway to crypt #1 is indicated as a cartoon.

Fig. 2.. Regulatory pathways controlling production of Syp-PS leading to biofilm formation and colonization.

As described in the text, the SKs RscS and HahK function upstream of the SK SypF, which in turn activates, via phosphorylation, the RR SypG. In turn, SypG activates transcription of the syp locus, resulting in production of the structural proteins necessary for Syp-PS production. Calcium is a positive signal inducing syp transcription that is inhibited by the SK BinK. Transcription is also inhibited by nitric oxide (NO) via HahK. Syp-PS production is further regulated post-transcriptionally by the action of the RR SypE, which phosphorylates and dephosphorylates SypA. Unphosphorylated SypA promotes Syp-PS production through an unknown mechanism.

Fig. 3.. Genetics and biochemistry of bioluminescence in V. fischeri.

a. Genetics and regulation: large arrows indicate the genes and their orientation at the lux locus. The bioluminescence reaction is driven by products of luxCDABEG, which are co-transcribed with luxI, and this “lux operon” is divergently transcribed from luxR. Two AHL signal molecules, N-3-oxohexanoyl homoserine lactone (3OC6), and N-octanoyl homoserine lactone (C8), produced by LuxI and AinS, respectively, are membrane permeable and can serve as cell-to-cell signals. When 3OC6 binds to LuxR, the activated regulator attaches to a sequence near the promoter of the lux operon called the “lux box”, resulting in an activation of transcription. While C8 is a weak activator of LuxR, it is strongly recognized by its cognate receptor, AinR, and works through a regulatory cascade to relieve the repression of LitR, a master regulator that controls transcription of several genes including LuxR. b. Biochemistry: the LuxAB heterodimer forms the luciferase enzyme, which sequentially binds reduced flavin mononucleotide (FMNH₂), O₂, and an aliphatic aldehyde (RCHO), and converts these substrates to FMN, water, and the corresponding aliphatic acid (RCOOH), emitting a bluish light (~490 nm wavelength) in the process. LuxC, LuxD, LuxE, and LuxG are responsible for (re)generating the substrates for luciferase.

Fig. 4.. The rise of V. fischeri genomic information.

Trajectory of the appearance of V. fischeri genome sequences over the last 15 years, which has seen a rapid increase since 2015. The geographical and biological sources of the isolates, as well as the number of strains sequenced, are indicated. Almost all the sequences are draft genomes. Associated publications are identified by their PMIDs: ^a15703294; ^b19182778; ^c22374964; ^d26044435; ^e(none); ^f27128997; ^g27653556; ^h31331977; ⁱ32127462.