Bacterial bioluminescence as a lure for marine zooplankton and fish Margarita Zarubina,b,1, Shimshon Belkinc, Michael Ionescuc, and Amatzia Genina,c aInteruniversity Institute for Marine Sciences, Eilat 88103, Israel; bInstitute for Chemistry and Biology of the Marine Environment, University of Oldenburg, 26111 Oldenburg, Germany; and cInstitute of Life Sciences, Hebrew University of Jerusalem, Jerusalem 91904, Israel Edited* by J. Woodland Hastings, Harvard University, Cambridge, MA, and approved December 2, 2011 (received for review October 11, 2011) The benefits of bioluminescence for nonsymbiotic marine bacteria zooplankton being involved, or by zooplankton that propagates have not been elucidated fully. One of the most commonly cited bacteria in its feces. explanations, proposed more than 30 y ago, is that biolumines- The objective of this study was to test the following key points cence augments the propagation and dispersal of bacteria by of the bait hypothesis: (i) visual attraction of zooplankton to attracting fish to consume the luminous material. This hypothesis, bacterial bioluminescence; (ii) promotion of glow in zooplankton based mostly on the prevalence of luminous bacteria in fish guts, contacting/ingesting luminous bacteria (using planktonic brine has not been tested experimentally. Here we show that zooplank- shrimps as a surrogate for zooplankton); (iii) attraction of zoo- ton that contacts and feeds on the luminescent bacterium Photo- planktivorous fish to glowing prey; and (iv) survival by bacteria of bacterium leiognathi starts to glow, and demonstrate by video gut passage in both zooplankton and fish. recordings that glowing individuals are highly vulnerable to pre- dation by nocturnal fish. Glowing bacteria thereby are transferred Results to the nutritious guts of fish and zooplankton, where they survive Zooplankton Attraction to Bacterial Bioluminescence. A large (135- digestion and gain effective means for growth and dispersal. L) experimental sea-water tank was used to examine whether the Using bioluminescence as bait appears to be highly beneficial luminescence of marine bacteria (Photobacterium leiognathi) for marine bacteria, especially in food-deprived environments of attracts zooplankton. A dialysis bag (20 mL) containing luminous the deep sea. bacteria, used as a bait, was placed at one corner of the tank; an ECOLOGY identical bag containing a culture of a dark mutant of P. leiog- ioluminescence is common in the marine environment, oc- nathi was placed at the opposite corner. Significant changes in Bcurring in numerous organisms, from bacteria to inverte- zooplankton distribution within the tank were noticeable within brates and fish (1, 2). Bacterial bioluminescence occurs as a 15 min. Decapods and mysids were found almost exclusively < continuous glow in the presence of oxygen at cell concentrations (one-sample t-test, P 0.001 for both decapods and mysids) over fi exceeding quorum-sensing levels (3–6). Luminous bacteria occur the glowing net, whereas copepods showed no signi cant at- traction (P = 0.269) to either net (Fig. 1). Similarly, no differ- free-living in seawater (7, 8), in symbiotic associations with ma- fi rine organisms (most notably fish and squids; see refs. 7 and 8 ence was found for nonmotile organisms (spherical sh eggs and Pyrocystis spp.; P = 0.34), which served as in internal control. and references therein), as saprophytes on suspended organic Together these four groups constituted on average 85% (range, material such as marine snow (9, 10), as a major component of 73–90%) of the plankton captured in our samples. Other taxo- fecal pellets (11–13), and as parasites on crustaceans (14). fi nomic groups were too rare to be included reliably in this Although the adaptive bene ts of energetically costly bio- comparison. luminescence in symbiotic bacteria are well understood (e.g., 7, fi 15), those bene ts in nonsymbiotic bacteria and those living as Zooplankton Turns Luminescent upon Contacting P. leiognathi ectoparasites on zooplankton are less obvious. Several different Cultures. The brine shrimp Artemia salina (hereafter “Artemia”) physiological and biochemical functions of bacterial bio- became luminescent after swimming for only 10 s in a liquid luminescence have been proposed (7, 16–20), focusing mostly on culture of P. leiognathi, as well as after swimming for 2.5 h in the antioxidative activity, enhanced DNA repair, and UV resistance, suspension of small particles of bioluminescent P. leiognathi although the validity of some of these hypotheses has been colonies. As revealed by long-exposure photographs, the lumi- questioned (21). nescence in the guts of Artemia was clearly visible from outside, An ecological function in propagation and dispersal also has with additional glow produced by bacteria attached externally to been postulated (6, 7, 22). According to this hypothesis (here- the exoskeleton and appendages (Fig. 2). Similarly, nonglowing after, “bait hypothesis“), the bacteria, by glowing, visually mark individual marine mysids, Anisomysis marisrubri, freshly trapped the presence of a food particle for fish in order to get into their in the sea, started to glow after contacting a diluted culture of nutritious guts. So far, this hypothesis was supported by cir- P. leiognathi. cumstantial evidence showing that luminous bacteria thrive in and survive passage through fish guts (7, 12, 23, 24). Here we Fish Detect and Consume Glowing Prey. The promoted glow in Artemia dramatically affected its risk of being preyed on by the propose that the mechanism underlying the bait hypothesis is fi based on the following steps: (i) Quorum sensing assures that nocturnal sh Apogon annularis in a recirculating laboratory fl Artemia bacterial bioluminescence is a reliable signal of the presence of ume in the dark. Almost all the glowing offered to the food aggregates, e.g., marine snow; (ii) zooplankton is attracted to luminous particles and grazes on the bacteria-rich organic Author contributions: M.Z., S.B., and A.G. designed research; M.Z. and M.I. performed matter; (iii) because of its contact with or ingestion of the lu- research; S.B. contributed new reagents/analytic tools; M.Z. and A.G. analyzed data; and minous bacteria, the zooplankton itself becomes glowing; (iv) the M.Z. and A.G. wrote the paper. glowing zooplankton is detected readily and consumed by fish; The authors declare no conflict of interest. (v) once in the gut of either zooplankton or fish, the bacteria gain *This Direct Submission article had a prearranged editor. a nutritious environment for growth and a fast-moving vehicle 1To whom correspondence should be addressed. [email protected]. fi for wide dispersal. The scheme may be shortened by sh that This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. directly detect and consume glowing organic particles without 1073/pnas.1116683109/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1116683109 PNAS Early Edition | 1of5 Downloaded by guest on September 23, 2021 1 * 0.8 * 0.6 0.4 0.2 0 Proportion in “glowing” net Fig. 2. Glow of zooplankton (A. salina) after contacting and ingesting small Mysida particles broken off colonies of the bioluminescent bacterium P. leiognathi. CopepodaFish eggs + Decapoda Pyrocystis spp. The photograph on the left was taken in room light, and the photograph on the right was taken in darkness using long exposure (30 s) with a Nikon D3 Fig. 1. Zooplankton attraction to the bioluminescence of P. leiognathi camera (f/5.6, ISO 25600, 150 mm lens). (Scale bar: 1 cm.) shown as average proportions (total in glowing net divided by totals in both nets + SEM; n = 8) of four selected zooplankton taxonomic groups. Asterisks indicate a significant difference (one-sample t test, P < 0.001 for each taxon) The next prediction of the bait hypothesis is that, on contact from the value of 0.5 expected under no attraction to either net (dotted line). with and ingestion of luminous particles, the zooplankton itself starts to glow, thereby attracting its own predators, such as fish. The advantage to the bacteria is obvious: By surviving digestion fish were consumed readily (Fig. 3 and Movie S1), compared in the guts of both zooplankton and fish, the bacteria gain a nu- with rare occasions of predation on nonglowing specimens. As trient-rich, sheltered environment for proliferation as well as an the video recordings revealed, the predation on nonglowing efficient means of dispersal. The advantage for zooplankton is specimens occurred only when the prey drifted by chance directly less obvious because of the tradeoff between the gain provided toward the fish’s head. The effect of glow of the prey on fish by organic-rich food and the cost incurred by a higher risk predation was highly significant (two-way ANOVA, F = 1,18 of predation. 275.648, P < 0.0001), whereas the effect of the identity of the fish To examine the bait hypothesis, we experimentally tested its was not significant (two-way ANOVA, F = 0.990, P = 0.39). 2,18 key steps. First, we documented the visual attraction of marine Analysis of the video recordings revealed that the fish actively zooplankton to bacterial bioluminescence (Fig. 1). Biolumines- attacked and consumed glowing prey (Movie S1), whereas the ∼ nonglowing prey passed undetected even at very close proximity cence in most marine bacteria peaks at the wavelength of 490 to the fish (Movie S2). Glowing prey were detected by the fish nm (25), which, not surprisingly, is near the wavelength least from a distance of up to 26.8 cm, near the limit allowed by the absorbed in seawater (26). Several zooplankton taxa [e.g., two working section of the flume. In fact, video records revealed that species of the copepod Pleuromamma (27), hyperiid amphi- the fish occasionally swam to the upstream part of the working pods (28), and some deep-sea crustaceans (29)] were shown to be sensitive to similar wavelengths. Mesopelagic crustaceans section and seemed to wait for the prey to arrive.