Dietary maintenance of bioluminescence in a deep-sea mysid

A major difficulty in studying dietary bioluminescence induction in marine organisms is finding experimental subjects that survive under strict dietary control for extended periods. The robust mysid shrimp, Gnathophausia ingens (Dohrn), has proved to be ideal for such studies. Relatively abundant off the coast of Southern California, it survives in the laboratory for up to 2·5 years (Childress & Price, 1978). Mechanically stimulated G. ingens secrete bluish luminescent material (Fig. 1) from a gland opening on each second maxilla at the base of the exognaths (Illig, 1905). Laboratory maintained specimens usually became non-luminescent after 2 months, but some retained the ability to luminesce for up to 8 months. We have now demonstrated that G. ingens which have lost luminescence capacity on a diet restricted to tissues from non-bioluminescent animals rapidly regain this capacity after ingesting certain luminescent prey.

Specimens of G. ingens, trawled at depths of 600–800 m in San Clemente Basin (off the coast of southern California), during February and August, 1982, were maintained separately in one-quart containers of sea water at 5 °C (Childress & Price, 1978). All were fed mackerel or salmon and tested for bioluminescence once a week. Most were no longer luminescent 4 months after capture, and all the animals used in dietary induction tests had not luminesced in response to mechanical stimulation for at least 1 month before experiments began.

Non-luminescent G. ingens were fed seven types of luminescent food items, namely : (1) euphausiid shrimps with photophores (live Euphaùsia pacifica and Nematoscelis difficilis); (2) a gonostomatid fish with photophores (live Cyclothone acclinidens) ; (3) a myctophid fish with photophores and caudal luminescent organs (pieces of freshly killed Triphoturus mexicanas) ; (4) a batrachoidid fish with photophores (pieces of freshly killed Porichthys notatus); (5) a luminescent coelenterate (frozen Atolla sp.); (6) a copepod possessing glands which release luminescent fluid (frozen Gaussia princeps) and (7) a shrimp possessing internal hepatic light organs (live Sergestes similis). They were fed every 2–3 days and tested for bioluminescence the day after each feeding, with the following results: 36 of 41 experimental animals regained luminescence capability after ingesting Triphoturus, Cyclothone, Atolla, Sergestes or Gaussia (Table 1). Three animals in these five groups never regained their luminescence, even though two of them received two different luminescent food items. Subsequent dissection showed that the luminescent organs were intact. Twenty-six specimens were fed euphausiid shrimp or Porichthys, and all remained non-luminescent. Eleven of these non-luminescent individuals were subsequently fed Triphoturus, and became luminescent within 2 days, demonstrating that they were potentially able to manufacture a luminescent secretion, but not from the euphausiid or Porichthys food items.

At least three types of marine bioluminescent systems are known: coelenterate, dinoflagellate and Cypridina. They differ from one another in the chemical structure of their luciferins and luciferases, and by weak or absent in vitro cross-reactions between the components of the different systems. Prey representing these three systems were utilized in this investigation. While the luminescent system of the myctophid Triphoturus has not been analysed, Inoue, Okada, Kakoi & Goto (1977) and Inoue, Kakoi, Okada & Goto (1979) present evidence for coelenterate-type luciferin in one other myctophid, and Shimomura et al. (1980) confirm its presence in two myctophid species. Although some degree of cross-reactivity with Cypridina luciferin may occur with impure extracts (Tsuji & Haneda, 1971), it is likely that the bioluminescent system of myctophids utilizes coelenterate-type luciferin (Cormier, 1978). Sergestes similis probably possesses coelenterate-type luciferin since it is present in two other Sergestes species (Shimomura et al. 1980). The luminescent system of the copepod, Gaus si a princeps, has not been characterized, but the luciferin of Pleuromamma pisecki, a copepod belonging to the same family, has been categorized as coelenterate-type (McCapra & Hart, 1980), supporting the possibility of coelenterate-type luciferin in Gaussia. Euphausiids apparently possess a different luminescent system. Even though aqueous extracts of the luminescent system of the euphausiid, Euphausia similis, give luminescent cross-reactions with crude Cypridina luciferase (Tsuji et al. 1971), other studies report that euphausiids have a photoprotein system unlike the luciferin/luciferase system of Cypridina (Shimomura & Johnson, 1967, 1968), and more recently, euphausiid fluorescent substance has been shown to be biochemically similar to dinoflagellate luciferin (Dunlap, Hastings & Shimomura, 1980). The luminescent system of Porichthys notatus is of the Cypridina type (Cormier et al. 1967; Tsuji et al. 1971, 1975). The luminescent system of Gnathophausia longispina is characterized as coelenterate-type (Shimomura et al. 1980) and the luciferin of G. ingens is related to the coelenteratetype (Cormier, 1978). It is apparent from the present study that G. ingens is able to utilize the components of coelenterate-type luminescent systems in myctophids and sergestids, but not the dinoflagellate-type system found in euphausiids, or the Cypridina-type system of Porichthys. Although the luminescent systems of Cyclothone and Atolla have not been analysed, this study strongly suggests that they possess coelenterate-type luminescence, since Gnathophausia can utilize their systems. Since the luminescent system of Gnathophausia appears specific for coelenterate-type luciferin/lucifcrase, it may be useful as a bioassay for animals possessing coelenterate-type luminescence. Studies are in progress to characterize more fully the specificity of the Gnathophausia luminescent system.

Luminescent emission spectra were determined for Triphoturus, freshly trawled G. ingens, and G. ingens with luminescent capability restored after ingesting Triphoturus. Spectra taken with an optical multichannel analyser (Princeton Applied Research OMA II) were standardized and digitally smoothed (Widder, Latz & Case, 1983). The Triphoturus spectrum (caudal organ) differs substantially from that of G. ingens, with the spectral peak shifted 12 nm towards the shorter wavelengths, and with a markedly more narrow full width at half maximum intensity (FWHM), 62 nm compared to 83 nm for Gnathophausia (Fig. 2A). Natural and Tnphoturus-’mduccd G. ingens spectra have identical λ_max at 481 nm and the same FWHM (Fig. 2B).

The differences in luminescence spectra between G. ingens and Triphoturus argue that some aspect of the chemistry of the two luminescent mechanisms may differ. A similar situation exists in the Porichthys-Cypridina in vivo interaction in which it is known that the enzyme originates with Porichthys and the Porichthys emission is bimodal (485 and 507 nm) while Cypridina luciferin and luciferase emit light with a single peak at 459nm (Reynolds, Botos & Barba, 1972; Tsuji et al. 1975). The variation in emission spectra may be caused by other factors, however, such as the reaction environment, which is intracellular in Triphoturus and extracellular in G. ingens.

Since some coelenterate-type bioluminescence is calcium dependent (Cormier, 1978; Shimomura, Johnson & Saiga, 1962), calcium might be expected to play a role in the luminescence of Gnathophausia. However, preliminary experiments indicate that the Gnathophausia luminescent system is not dependent on external calcium, since luminescence appears normal when animals are maintained for 12 h in calcium-free, artificial sea water containing 10 mg 1⁻¹ EGTA and caused to secrete into the same or into distilled water.

We thank Drs J. J. Childress and A. J. Arp for help with the collection of these organ isms. Supported by U.S. Office of Naval Research Contract N00014-75-C-0242.