Ultrasensitive and robust mechanoluminescent living composites.

Mechanosensing, the transduction of extracellular mechanical stimuli into intracellular biochemical signals, is a fundamental property of living cells. However, endowing synthetic materials with mechanosensing capabilities comparable to biological levels is challenging. Here, we developed ultrasensitive and robust mechanoluminescent living composites using hydrogels embedded with dinoflagellates, unicellular microalgae with a near-instantaneous and ultrasensitive bioluminescent response to mechanical stress. Not only did embedded dinoflagellates retain their intrinsic mechanoluminescence, but with hydrophobic coatings, living composites had a lifetime of ~5 months under harsh conditions with minimal maintenance. We 3D-printed living composites into large-scale mechanoluminescent structures with high spatial resolution, and we also enhanced their mechanical properties with double-network hydrogels. We propose a counterpart mathematical model that captured experimental mechanoluminescent observations to predict mechanoluminescence based on deformation and applied stress. We also demonstrated the use of the mechanosensing composites for biomimetic soft actuators that emitted colored light upon magnetic actuation. These mechanosensing composites have substantial potential in biohybrid sensors and robotics.

Highly robust and soft biohybrid mechanoluminescence for optical signaling and illumination.

Biohybrid is a newly emerging and promising approach to construct soft robotics and soft machines with novel functions, high energy efficiency, great adaptivity and intelligence. Despite many unique advantages of biohybrid systems, it is well known that most biohybrid systems have a relatively short lifetime, require complex fabrication process, and only remain functional with careful maintenance. Herein, we introduce a simple method to create a highly robust and power-free soft biohybrid mechanoluminescence, by encapsulating dinoflagellates, bioluminescent unicellular marine algae, into soft elastomeric chambers. The dinoflagellates retain their intrinsic bioluminescence, which is a near-instantaneous light response to mechanical forces. We demonstrate the robustness of various geometries of biohybrid mechanoluminescent devices, as well as potential applications such as visualizing external mechanical perturbations, deformation-induced illumination, and optical signaling in a dark environment. Our biohybrid mechanoluminescent devices are ultra-sensitive with fast response time and can maintain their light emission capability for weeks without special maintenance.

Molecular and biochemical basis for the loss of bioluminescence in the dinoflagellate Noctiluca scintillans along the west coast of the USA.

The globally distributed heterotrophic dinoflagellate Noctiluca scintillans (Macartney) Kofoid & Swezy is well known for its dense blooms and prominent displays of bioluminescence. Intriguingly, along the west coast of the USA its blooms are not bioluminescent. We investigated the basis for the regional loss of bioluminescence using molecular, cellular and biochemical analyses of isolates from different geographic regions. Prominent differences of the non-bioluminescent strains were: (1) the fused luciferase and luciferin binding protein gene (lcf/lbp) was present but its transcripts were undetectable; (2) lcf/lbp contained multiple potentially deleterious mutations; (3) the substrate luciferin was absent, based on the lack of luciferin blue autofluorescence and the absence of luciferin derived metabolites; (4) although the cells possessed scintillons, the vesicles that contain the luminescent chemistry, electron microscopy revealed additional scintillon-like vesicles with an atypical internal structure; (5) cells isolated from the California coast were 43% smaller in size than bioluminescent cells from the Gulf of Mexico. Phylogenetic analyses based on the large subunit of rDNA did not show divergence of the non-bioluminescent population in relation to other bioluminescent N. scintillans from the Pacific Ocean and Arabian Sea. Our study demonstrates that gene silencing and the lack of the luciferin substrate have resulted in the loss of a significant dinoflagellate functional trait over large spatial scales in the ocean. As the bioluminescence system of dinoflagellates is well characterized, non-bioluminescent N. scintillans is an ideal model to explore the evolutionary and ecological mechanisms that lead to intraspecific functional divergence in natural dinoflagellate populations.

Bubble stimulation efficiency of dinoflagellate bioluminescence.

Dinoflagellate bioluminescence, a common source of bioluminescence in coastal waters, is stimulated by flow agitation. Although bubbles are anecdotally known to be stimulatory, the process has never been experimentally investigated. This study quantified the flash response of the bioluminescent dinoflagellate Lingulodinium polyedrum to stimulation by bubbles rising through still seawater. Cells were stimulated by isolated bubbles of 0.3-3 mm radii rising at their terminal velocity, and also by bubble clouds containing bubbles of 0.06-10 mm radii for different air flow rates. Stimulation efficiency, the proportion of cells producing a flash within the volume of water swept out by a rising bubble, decreased with decreasing bubble radius for radii less than approximately 1 mm. Bubbles smaller than a critical radius in the range 0.275-0.325 mm did not stimulate a flash response. The fraction of cells stimulated by bubble clouds was proportional to the volume of air in the bubble cloud, with lower stimulation levels observed for clouds with smaller bubbles. An empirical model for bubble cloud stimulation based on the isolated bubble observations successfully reproduced the observed stimulation by bubble clouds for low air flow rates. High air flow rates stimulated more light emission than expected, presumably because of additional fluid shear stress associated with collective buoyancy effects generated by the high air fraction bubble cloud. These results are relevant to bioluminescence stimulation by bubbles in two-phase flows, such as in ship wakes, breaking waves, and sparged bioreactors.

Mechanosensitivity of a rapid bioluminescence reporter system assessed by atomic force microscopy.

Cells are sophisticated integrators of mechanical stimuli that lead to physiological, biochemical, and genetic responses. The bioluminescence of dinoflagellates, alveolate protists that use light emission for predator defense, serves as a rapid noninvasive whole-cell reporter of mechanosensitivity. In this study, we used atomic force microscopy (AFM) to explore the relationship between cell mechanical properties and mechanosensitivity in live cells of the dinoflagellate Pyrocystis lunula. Cell stiffness was 0.56 MPa, consistent with cells possessing a cell wall. Cell response depended on both the magnitude and velocity of the applied force. At the maximum stimulation velocity of 390 µm s(-1), the threshold response occurred at a force of 7.2 µN, resulting in a contact time of 6.1 ms and indentation of 2.1 µm. Cells did not respond to a low stimulation velocity of 20 µm s(-1), indicating a velocity dependent response that, based on stress relaxation experiments, was explained by the cell viscoelastic properties. This study demonstrates the use of AFM to study mechanosensitivity in a cell system that responds at fast timescales, and provides insights into how viscoelastic properties affect mechanosensitivity. It also provides a comparison with previous studies using hydrodynamic stimulation, showing the discrepancy in cell response between direct compressive forces using AFM and those within flow fields based on average flow properties.

Pharmacological investigation of the bioluminescence signaling pathway of the dinoflagellate Lingulodinium polyedrum: evidence for the role of stretch-activated ion channels.

Dinoflagellate bioluminescence serves as a whole-cell reporter of mechanical stress, which activates a signaling pathway that appears to involve the opening of voltage-sensitive ion channels and release of calcium from intracellular stores. However, little else is known about the initial signaling events that facilitate the transduction of mechanical stimuli. In the present study using the red tide dinoflagellate Lingulodinium polyedrum (Stein) Dodge, two forms of dinoflagellate bioluminescence, mechanically stimulated and spontaneous flashes, were used as reporter systems to pharmacological treatments that targeted various predicted signaling events at the plasma membrane level of the signaling pathway. Pretreatment with 200 µM Gadolinium III (Gd(3+) ), a nonspecific blocker of stretch-activated and some voltage-gated ion channels, resulted in strong inhibition of both forms of bioluminescence. Pretreatment with 50 µM nifedipine, an inhibitor of L-type voltage-gated Ca(2+) channels that inhibits mechanically stimulated bioluminescence, did not inhibit spontaneous bioluminescence. Treatment with 1 mM benzyl alcohol, a membrane fluidizer, was very effective in stimulating bioluminescence. Benzyl alcohol-stimulated bioluminescence was inhibited by Gd(3+) but not by nifedipine, suggesting that its role is through stretch activation via a change in plasma membrane fluidity. These results are consistent with the presence of stretch-activated and voltage-gated ion channels in the bioluminescence mechanotransduction signaling pathway, with spontaneous flashing associated with a stretch-activated component at the plasma membrane.

Green fluorescent protein regulation in the coral Acropora yongei during photoacclimation.

Reef-building corals inhabit high light environments and are dependent on photosynthetic endosymbiotic dinoflagellates for nutrition. While photoacclimation responses of the dinoflagellates to changes in illumination are well understood, host photoacclimation strategies are poorly known. This study investigated fluorescent protein expression in the shallow-water coral Acropora yongei during a 30 day laboratory photoacclimation experiment in the context of its dinoflagellate symbionts. Green fluorescent protein (GFP) concentration measured by Western blotting changed reversibly with light intensity. The first 15 days of the photoacclimation experiment led to a ∼1.6 times increase in GFP concentration for high light corals (900 µmol quanta m⁻² s⁻¹) and a ∼4 times decrease in GFP concentration for low light corals (30 µmol quanta m⁻² s⁻¹) compared with medium light corals (300 µmol quanta m⁻² s⁻¹). Green fluorescence increased ∼1.9 times in high light corals and decreased ∼1.9 times in low light corals compared with medium light corals. GFP concentration and green fluorescence intensity were significantly correlated. Typical photoacclimation responses in the dinoflagellates were observed including changes in density, photosynthetic pigment concentration and photosynthetic efficiency. Although fluorescent proteins are ubiquitous and abundant in scleractinian corals, their functions remain ambiguous. These results suggest that scleractinian corals regulate GFP to modulate the internal light environment and support the hypothesis that GFP has a photoprotective function. The success of photoprotection and photoacclimation strategies, in addition to stress responses, will be critical to the fate of scleractinian corals exposed to climate change and other stressors.

Bioluminescent response of individual dinoflagellate cells to hydrodynamic stress measured with millisecond resolution in a microfluidic device.

Dinoflagellate bioluminescence serves as a model system for examining mechanosensing by suspended motile unicellular organisms. The response latency, i.e. the delay time between the mechanical stimulus and luminescent response, provides information about the mechanotransduction and signaling process, and must be accurately known for dinoflagellate bioluminescence to be used as a flow visualization tool. This study used a novel microfluidic device to measure the response latency of a large number of individual dinoflagellates with a resolution of a few milliseconds. Suspended cells of several dinoflagellate species approximately 35 microm in diameter were directed through a 200 microm deep channel to a barrier with a 15 microm clearance impassable to the cells. Bioluminescence was stimulated when cells encountered the barrier and experienced an abrupt increase in hydrodynamic drag, and was imaged using high numerical aperture optics and a high-speed low-light video system. The average response latency for Lingulodinium polyedrum strain HJ was 15 ms (N>300 cells) at the three highest flow rates tested, with a minimum latency of 12 ms. Cells produced multiple flashes with an interval as short as 5 ms between individual flashes, suggesting that repeat stimulation involved a subset of the entire intracellular signaling pathway. The mean response latency for the dinoflagellates Pyrodinium bahamense, Alexandrium monilatum and older and newer isolates of L. polyedrum ranged from 15 to 22 ms, similar to the latencies previously determined for larger dinoflagellates with different morphologies, possibly reflecting optimization of dinoflagellate bioluminescence as a rapid anti-predation behavior.

Bioluminescence characteristics of a tropical terrestrial fungus (Basidiomycetes).

Freshly collected samples of luminous mycelium of a terrestrial fungus from Panama were investigated for their bioluminescence characteristics. Taxonomic identification of fungal species could not be determined because of the lack of fruiting bodies. Fluorescence excited by 380 nm illumination had an emission spectrum with a main peak at 480 nm and additional chlorophyll peaks related to the wood substrate. Bioluminescence appeared as a continuous glow that did not show any diel variation. The light production was sensitive to temperature and decreased with temperatures higher or lower than ambient. Bioluminescence intensity was sensitive to hydration, increasing by a factor of 400 immediately after exposure to water and increasing by a factor of 1 million after several hours. This increase may have occurred through dilution of superoxide dismutase, which is a suppressive factor of bioluminescence in fungus tissue. The mycelium typically transports nutritive substances back to the fruiting body. The function of luminescent mycelium may be to increase the intensity of light from the fungus and more effectively attract nocturnal insects and other animals that serve as disseminating vectors for fungal spores.

Shear-stress dependence of dinoflagellate bioluminescence.

Fluid flow stimulates bioluminescence in dinoflagellates. However, many aspects of the cellular mechanotransduction are incompletely known. The objective of our study was to formally test the hypothesis that flow-stimulated dinoflagellate bioluminescence is dependent on shear stress, signifying that organisms are responding to the applied fluid force. The dinoflagellate Lingulodinium polyedrum was exposed to steady shear using simple Couette flow in which fluid viscosity was manipulated to alter shear stress. At a constant shear rate, a higher shear stress due to increased viscosity increased both bioluminescence intensity and decay rate, supporting our hypothesis that bioluminescence is shear-stress dependent. Although the flow response of non-marine attached cells is known to be mediated through shear stress, our results indicate that suspended cells such as dinoflagellates also sense and respond to shear stress. Shear-stress dependence of flow-stimulated bioluminescence in dinoflagellates is consistent with mechanical stimulation due to direct predator handling in the context of predator-prey interactions.

Evidence for the role of G-proteins in flow stimulation of dinoflagellate bioluminescence.

Luminescent dinoflagellates respond to flow by the production of light. The primary mechanotransduction event is unknown, although downstream events include a calcium flux in the cytoplasm, a self-propagating action potential across the vacuole membrane, and a proton flux into the cytoplasm that activates the luminescent chemistry. Given the role of GTP-binding (G) proteins in the mechanotransduction of flow by nonmarine cells and the presence of G-proteins in dinoflagellates, it was hypothesized that flow-stimulated dinoflagellate bioluminescence involves mechanotransduction by G-proteins. In the present study, osmotic swelling of cells of the dinoflagellate Lingulodinium polyedrum was used as a drug delivery system to introduce GDPbetaS, an inhibitor of G-protein activation. Osmotically swollen cells produced higher levels of flow-stimulated bioluminescence at a lower threshold of shear stress, indicating they were more flow sensitive. GDPbetaS inhibited flow-stimulated bioluminescence in osmotically swollen cells and in cells that were restored to the isosmotic condition following hypoosmotic treatment with GDPbetaS. These results provide evidence that G-proteins are involved in the mechanotransduction of flow in dinoflagellates and suggest that G-protein involvement in mechanotransduction may be a fundamental evolutionary adaptation.

Bioavailability of metals along a contamination gradient in San Diego Bay (California, USA).

San Diego Bay is heavily contaminated with metals, but little is known about their biological availability to local marine organisms. This study on 15 elements showed that concentrations of metals associated with sediment increased from the mouth to the back of the Bay while metals in seawater particulates were similar throughout the Bay. Metal bioavailability was assessed over 8weeks following transplant of the local brittlestar, Ophiothrix spiculata (Ophuroidea, Echinodermata), from outside to inside the Bay. Despite a gradient of contamination, brittlestars accumulated similar levels of metals throughout the Bay, suggesting that metal contamination occurred through dissolved metals as well as through the diet. Sediment transplanted in dialysis tubing in the Bay accumulated metals only when placed on the seafloor bottom, indicating greater metal bioavailability near the bottom; the level of accumulation was similar between the mouth and the back of the Bay. The results are consistent with a circulation pattern in which a bottom layer of seawater, enriched with metals, drains from the back to the mouth of the Bay. There was a positive correlation between metal concentration in brittlestars and tidal range, suggesting increased metal exposure due to bay-ocean water exchange. For brittlestar arms the correlation was higher at the mouth than the back of the Bay, indicating greater metal accumulation in arms from dissolved metals in seawater than from ingestion of metal contaminated diet. In contrast, for brittlestar disks the correlation was higher at the back of the Bay, indicative of metal accumulation mainly through the diet. The results highlight the importance of considering bioavailability and physical processes in environmental quality assessments.

Evidence for enhanced bioavailability of trace elements in the marine ecosystem of Deception Island, a volcano in Antarctica.

This study assessed whether trace elements present at Deception Island, an active submarine volcano in the Antarctic Peninsula, show enhanced biological availability to the local marine community. Using a weak acid extraction method to dissolve organic material and leach associated but not constitutive trace elements of sediments, fifteen elements were measured from seafloor sediment, seawater particulates, and tissues of benthic (bivalves, brittlestars, sea urchins) and pelagic (demersal and pelagic fishes, krill) organisms collected in the flooded caldera. The highest element concentrations were associated with seafloor sediment, the lowest with seawater particulates and organism tissues. In the case of Ag and Se, concentrations were highest in organism tissue, indicating contamination through the food chain and biomagnification of those elements. The elements Al, Fe, Mn, Sr, Ti, and to a lesser extent Zn, were the most concentrated of the trace elements for all sample types. This indicates that the whole ecosystem of Deception Island is contaminated with trace elements from local geothermal activity, which is also reflected in the pattern of element contamination in organisms. Accordingly, element concentrations were higher in organisms collected at Deception Island compared to those from the neighboring non-active volcanic King George Island, suggesting that volcanic activity enhances bioavailability of trace elements to marine organisms. Trace element concentrations were highest in digestive tissue of organisms, suggesting that elements at Deception Island are incorporated into the marine food web mainly through a dietary route.

Chemical speciation and toxicity of metals assessed by three bioluminescence-based assays using marine organisms.

Metal toxicity is a function of the biology of the target organism and the chemical speciation of the metal. The toxicity of 11 metals was assessed with three cell-based bioassays based on marine organisms: the bacterium Photobacterium phosphoreum of the Microtox bioassay, an environmental strain of P. phosphoreum, and photocytes isolated from the brittlestar Ophiopsila californica. Metal speciation was calculated for three commonly used media: NaCl-based Microtox bioassay medium, artificial seawater glycerol, and artificial seawater. Decreased bioluminescence was considered a proxy for cell toxicity. In all three assays the elements Cd and Hg exhibited similar speciation as well as similar toxicity profiles. The element Cu was toxic in all three assays despite different metal speciation for the P. phosphoreum bioassay. The element Ag was toxic to both bacterial strains but not to photocytes despite a similar chemical speciation for all three assays. In general, the Microtox bioassay was sensitive to all metals (except Pb), whereas the photocytes were the least sensitive to the metals. The heightened response of the Microtox bioassay probably resulted from a combination of the limited complexing power of the medium and the greater sensitivity of the bacterial strain.

Hydrodynamic stimulation of dinoflagellate bioluminescence: a computational and experimental study.

Dinoflagellate bioluminescence provides a near-instantaneous reporter of cell response to flow. Although both fluid shear stress and acceleration are thought to be stimulatory, previous studies have used flow fields dominated by shear. In the present study, computational and experimental approaches were used to assess the relative contributions to bioluminescence stimulation of shear stress and acceleration in a laminar converging nozzle. This flow field is characterized by separate regions of pronounced acceleration away from the walls, and shear along the wall. Bioluminescence of the dinoflagellates Lingulodinium polyedrum and Ceratocorys horrida, chosen because of their previously characterized different flow sensitivities, was imaged with a low-light video system. Numerical simulations were used to calculate the position of stimulated cells and the levels of acceleration and shear stress at these positions. Cells were stimulated at the nozzle throat within the wall boundary layer where, for that downstream position, shear stress was relatively high and acceleration relatively low. Cells of C. horrida were always stimulated significantly higher in the flow field than cells of L. polyedrum and at lower flow rates, consistent with their greater flow sensitivity. For both species, shear stress levels at the position of stimulated cells were similar to but slightly greater than previously determined response thresholds using independent flow fields. L. polyedrum did not respond in conditions where acceleration was as high as 20 g. These results indicate that shear stress, rather than acceleration, was the stimulatory component of flow. Thus, even in conditions of high acceleration, dinoflagellate bioluminescence is an effective marker of shear stress.

The use of dinoflagellate bioluminescence to characterize cell stimulation in bioreactors.

Bioluminescent dinoflagellates are flow-sensitive marine organisms that produce light emission almost instantaneously upon stimulation by fluid shear in a shear stress dose-dependent manner. In the present study we tested the hypothesis that monitoring bioluminescence by suspended dinoflagellates can be used as a tool to characterize cellular response to hydrodynamic forces in agitated bioreactors. Specific studies were performed to determine: (1) impeller configurations with minimum cell activation, (2) correlations of cellular response and an integrated shear factor, and (3) the effect of rapid acceleration in agitation. Results indicated that (1) at a volumetric mass transfer coefficient of 3 x 10(-4) s(-1), marine impeller configurations were less stimulatory than Rushton configurations, (2) bioluminescence response and a modified volumetric integrated shear factor had an excellent correlation, and (3) rapid acceleration in agitation was highly stimulatory, suggesting a profound effect of temporal gradients in shear in increasing cell stimulation. By using bioluminescence stimulation as an indicator of agitation-induced cell stimulation and/or damage in microcarrier cultures, the present study allows for the verification of hypotheses and development of novel mechanisms of cell damage in bioreactors.

The role of Ca(2+) in stimulated bioluminescence of the dinoflagellate Lingulodinium polyedrum.

Many marine dinoflagellates emit bright discrete flashes of light nearly instantaneously in response to either laminar or turbulent flows as well as to direct mechanical stimulation. The flash involves a unique pH-dependent luciferase and a proton-mediated action potential across the vacuole membrane. The mechanotransduction process initiating this action potential is unknown. The present study investigated the role of Ca(2+) in the mechanotransduction process regulating bioluminescence in the dinoflagellate Lingulodinium polyedrum. Calcium ionophores and digitonin stimulated luminescence in a Ca(2+)-dependent manner in the absence of mechanical stimulation. Mechanically sensitive luminescence was strongly inhibited by the intracellular Ca(2+) chelator BAPTA-AM [1,2-bis(o-aminophenoxy)ethane-N,N,N',N'- tetraacetic acid acetoxymethyl ester]; there was only a partial and irreversible dependence on extracellular Ca(2+). Ruthenium Red, a blocker of intracellular Ca(2+) release channels, inhibited mechanically sensitive luminescence. Luminescence was also stimulated by increasing K(+), even in the absence of extracellular Ca(2+); K(+) stimulation was inhibited both by BAPTA-AM and Ruthenium Red. These results support the hypothesis that Ca(2+) mediates stimulated bioluminescence and also indicate the involvement of intracellular Ca(2+) stores. Rapid coupling between mechanical stimulation and mobilization of intracellular Ca(2+) stores might occur through a mechanism similar to excitation-contraction coupling in skeletal muscle.

VARIABILITY IN FLASH CHARACTERISTICS OF A BIOLUMINESCENT COPEPOD.

Bioluminescence of the copepod, Pleuromamma xiphias, was investigated with an optical multichannel analyzer(OMA) to measure emission spectra, an integrating sphere-photon counting detector system to determine flash kinetics and quantum emission, and an ISIT video system to image spatial patterns of emission. Light emission was in the blue spectral region, with maximum emission at approximately 492 nm. Spectral waveforms were unimodal, or bimodal with the secondary peak at 472 nm. Flashes in response to a single stimulus consisted of two components: a fast component attaining maximum intensity in under 100 ms, and a slow element which peaked after 600 ms. The fast component originated from thoracic and abdominal light organs while the slow component represented a large expulsion of luminescent material from the abdominal organ only. Both components exhibited first order exponential decay although the decay rate of the fast component was approximately one order of magnitude greater. The typical flash response to a single stimulus exhibited a response latency of 30 ms, initial rise time of 87 ms, duration of 2.4 s, and quantum emission of 1.4 x 1010 photons flash-1. Quantum emission increased with increasing stimulus strength. Both response waveform and total quantum emission were affected by the frequency of electrical stimuli. Stimulation at 1 Hz generated the greatest luminescence, averaging 1.1 x 1011 photons response-1 for 11 s emissions. Higher rates of stimulation decreased total quantum emission and response episode duration, and resulted in greater temporal summation of the emission waveform. Variability in flash characteristics due to electrical stimulation suggests a versatility of luminescent displays in situ.

MARINE BIOLUMINESCENCE SPECTRA MEASURED WITH AN OPTICAL MULTICHANNEL DETECTION SYSTEM.

The emission spectra of 70 bioluminescent marine species were measured with a computer controlled optical multichannel analyzer (OMA). A 350 nm spectral window is simultaneously measured using a linear array of 700 silicon photodiodes, coupled by fiber optics to a microchannel plate image intensifier on which a polychromator generated spectrum is focused. Collection optics include a quartz fiber optic bundle which allows spectra to be measured from single photophores. Since corrections are not required for temporal variations in emissions, it was possible to acquire spectra of transient luminescent events that would be difficult or impossible to record with conventional techniques. Use of this system at sea on freshly trawled material and in the laboratory has permitted acquisition of a large collection of bioluminescence spectra of precision rarely obtained previously with such material. Among unusual spectral features revealed were organisms capable of emitting more than one color, including: Umbellula magniflora and Stachyptilum superbum (Pennatulacea), Parazoanthus lucificum (Zoantharia), and Cleidopus gloria-maris (Pisces). Evidence is presented that the narrow bandwidth of the emission spectrum for Argyropelecus affinis (Pisces) is due to filters in the photophores.