The Luminescent Systems of Pony Fishes

The anatomy of bioluminescent organs and mode of light production in 18 species of pony fish have been investigated using fresh and preserved material. The luminescent systems are similarly arranged in all. Basically, the system consists of a light organ located at the distal end of the esophagus, and a series of abdominal accessory structures positioned in tandem for controlling light intensity and for directing and dispersing the light. Light is produced by numerous symbiotic luminous bacteria in the light organ. A simple classification of the luminescent systems is proposed. The light organs of Leiognathus elongatus and L. rivulatus show marked sexual dimorphism. The bacteria present in the light organs of many pony fishes are easily culturable, but not those from L. elongatus. Electron micrographs of the light organs of L. elongatus and L. rivulatus show the presence of numerous rod-shaped bacteria measuring approximately 0.8 µ x 2.4 µ and 0.8 µ x 7.3 µ, respectively. It is concluded that the light organ of L. elongatus contains another example of a type of non-culturable luminous bacteria that have been found elsewhere. Such bacteria appear to require from the host some special factor for growth and luminescence.     

Spectral characteristics of the bioluminescence induced in the marine fish,Porichthys notatus,byCypridina (ostracod) luciferin

Summary Specimens ofPorichthys notatus, which are naturally luminous along the coast of California, are non-luminous in Puget Sound. However, luminescence capability may be induced in the adult Puget SoundPorichthys by the administration of purifiedCypridina (ostracod) luciferin, syntheticCypridina luciferin, orCypridina organisms. The bioluminescence emission spectra produced by the Puget Sound fish following induction is similar, if not identical, to that of the naturally luminousPorichthys notatus from California waters (maxima: 485 and 507 nm).     

Contribution by symbiotically luminous fishes to the occurrence and bioluminescence of luminous bacteria in seawater

Seawater samples from a variety of locations contained viable luminous bacteria, but luminescence was not detectable although the system used to measure light was sensitive enough to measure light from a single, fully induced luminous bacterial cell. When the symbiotically luminous fishCleidopus gloriamaris was placed in a sterile aquarium, plate counts of water samples showed an increase in luminous colony-forming units. Luminescence also increased, decreasing when the fish was removed. Light measurements of water samples from a sterile aquarium containingPhotoblepharon palpebratus, another symbiotically luminous fish, whose bacterial symbionts have not been cultured, showed a similar pattern of increasing light which rapidly decreased upon removal of the fish. These experiments suggest that symbiotically luminous fishes release brightly luminous bacteria from light organs into their environment and may be a source of planktonic luminous bacteria. Although planktonic luminous bacteria are generally not bright when found in seawater, water samples from environments with populations of symbiotically luminous fish may show detectable levels of light.     

Bioluminescence in the symbiotic squid Euprymna scolopes is controlled by a daily biological rhythm

In most symbioses between animals and luminous bacteria it has been assumed that the bacterial symbionts luminesce continuously, and that the control of luminescent output by the animal is mediated through elaborate accessory structures, such as chromatophores and muscular shutters that surround the host light organ. However, we have found that while in the light organ of the sepiolid squid Euprymna scolopes, symbiotic cells of Vibrio fischeri do not produce a continuously uniform level of luminescence, but instead exhibit predictable cyclic fluctuations in the amount of light emitted per cell. This daily biological rhythm exhibits many features of a circadian pattern, and produces an elevated intensity of symbiont luminescence in juvenile animals during the hours preceding the onset of ambient darkness. Comparisons of the specific luminescence of bacteria in the intact light organ with that of newly released bacteria support the existence of a direct host regulation of the specific activity of symbiont luminescence that does not require the intervention of accessory tissues. A model encompassing the currently available evidence is proposed for the control of growth and luminescence activity in the E. scolopes/V. fischeri light organ symbiosis.Abbreviations CFU colony-forming-unit - LD light-dark     

Determination of the luciferin contents in luminous and non-luminous beetles

The contents of firefly luciferin in luminous and non-luminous beetles were determined by the methods of HPLC with fluorescence detection and the luminescence reaction of luciferin and firefly luciferase. Luminous cantharoids and elaterids contained various amounts of luciferin in the range of pmol to hundreds of nmol, but no luciferin was detected in the non-luminous cantharoids and elaterids.     

Determination of the Luciferin Contents in Luminous and Non-Luminous Beetles

The contents of firefly luciferin in luminous and non-luminous beetles were determined by the methods of HPLC with fluorescence detection and the luminescence reaction of luciferin and firefly luciferase. Luminous cantharoids and elaterids contained various amounts of luciferin in the range of pmol to hundreds of nmol, but no luciferin was detected in the non-luminous cantharoids and elaterids.     

Early development of bioluminescence suggests camouflage by counter‐illumination in the velvet belly lantern shark Etmopterus spinax (Squaloidea: Etmopteridae)

The development of luminous structures and the acquisition of luminescence competence during the ontogeny of the velvet belly lantern shark Etmopterus spinax, a deep‐sea squalid species, were investigated. The sequential appearance of nine different luminous zones during shark embryogenesis were established, and a new terminology for them given. These zones form the complex luminous pattern observed in free‐swimming animals. The organogenesis of photophores (photogenic organs) from the different luminous zones was followed, and photophore maturation was marked by the appearance of green fluorescent vesicles inside the photocytes (photogenic cells). Peroxide‐induced light emissions as well as spontaneous luminescence analysis indicated that the ability of E. spinax to produce light was linked to the presence of these fluorescent vesicles and occured prior to birth. The size of photogenic organs, as well as the percentage of ventral body surface area occupied by the luminous pattern and covered by photophores increased sharply during embryogenesis but remained relatively stable in free‐swimming animals. All these results strongly suggest camouflage by counter‐illumination in juvenile E. spinax.     

Effect of light intensity on the locomotor activity rhythm of Talitrus saltator (Montagu 1808) from Korba Beach

In this work, we investigate the locomotor behaviour of Talitrus saltator (Montagu 1808) for a population collected from the supralittoral zone of Korba beach. The locomotor activity rhythm was recorded for adult individuals during 10 summer days under continuous light with four different luminous intensities: 5 lux (N = 30), 35 lux (N = 30), 75 lux (N = 30) and 140 lux (N = 15). By the end of the experiments, 100% of the considered individuals were found alive under light intensities of 35 and 140 lux, whereas only 86 and 90% were found alive under light intensity of 5 and 75 lux, respectively. Furthermore, whatever the imposed luminous intensity is, actograms showed a clear drift to the right lengthening day after day the circadian period. Moreover, we found that by raising the light intensity, the drift becomes increasingly important. Actograms as well as activity curves, results showed that the locomotor activity profiles are mainly unimodal and their percentage increases when increasing the light intensity. Furthermore, periodogram analysis highlighted the presence of ultradian and circadian components where the longest periods were observed with the highest luminous intensity. In addition, the locomotor activity rhythm was statistically more defined and individuals of Talitrus saltator were significantly more active under the lowest luminous intensity.     

Evidence of luminous bacterial symbionts in the light organs of myctophid and stomiiform fishes

The myctophids and stomiiforms represent two common groups of luminous fishes, but the source of luminescence in these animals has remained undetermined. In this study, labeled luciferase gene fragments from luminous marine bacteria were used to probe DNA isolated from specific fish tissues. A positive signal was obtained from skin DNA in all luminous fishes examined, whereas muscle DNA gave a weaker signal and brain DNA was negative. This observation is consistent with luminous bacteria acting as the light source in myctophids and stomiiforms and argues against the genes necessary for luminescence residing on the fish chromosomes. To confirm the location of this signal, a bacterial probe was hybridized in situ to sections of a stomiiform. A strong signal was generated directly over specific regions of the fish light organs, whereas no signal was found over other internal or epidermal tissues of the fish. Taken together, these data provide the first indication that luminous bacterial symbionts exist in myctophids and stomiiforms and that these symbionts account for luminescence in these fishes.     

Ontogeny of photophore pattern in the velvet belly lantern shark,Etmopterus spinax

Bioluminescence is known to be of great ecological importance to a luminous organism but extremely few studies investigate the ontogeny of luminous capabilities. The photogenic pattern of the velvet belly lantern shark Etmopterus spinax was investigated over ontogeny (14.0–52.5 cm total length) to determine the scaling of the surface area and the photophore density of different luminous zones as well as the ecological consequences of ontogenetic variations in bioluminescence efficiency. According to the luminous zone considered, different scaling patterns were found for the surface areas while the photophore densities of all zones scale with negative allometry, even though photophore insertion occurs. No sexual differences in these relationships were found. Luminous zones can be placed in two morphologically different groups: the “coverage” and the “isolated” zones. While counter-illumination is certainly the function of the former, the latter are probably involved in intraspecific behaviours. Due to the discrepancy between luminous capabilities of these two luminous zone categories, there is an ontogenetic increase in the luminescence heterogeneity of the luminous pattern as it was shown by luminescence modelling and confirmed by direct observations of spontaneous luminescence in living sharks. This heterogeneity certainly represents a trade-off between an efficient ventral camouflage and a strong identification tool for intraspecific behaviours such as coordinate hunting, which would be particularly useful when E. spinax become fish eaters (>19 cm total length), and for sexual recognition in mature individuals.     

Localization of the bioluminescence system in the pileus of Mycena chlorophos

Mycena chlorophos, which is primarily distributed in Southeast Asia, is a luminous fungus that emits a bright green light from its pileus for about 2 days at approximately 20°C and high relative humidity. The distribution of bioluminescent tissues in the whole pileus and its sections was heterogeneous. The light intensity in the cap and the upper region of the gill was greater than that in the lower region of the gill. At the microscopic level, the light was predominantly emitted from the membranes of hymenium and basidia cells on the gill. The emission was both cell and region specific. The luminescence system was localized in the cell membrane, and a part of the system was on the cell membrane surface. Copyright © 2015 John Wiley & Sons, Ltd.     

LABORATORY STUDIES OF A GREEN NOCTILUCA FROM NEW GUINEA1,2

A strain of Noctiluca miliaris Suriray, containing in the vacuole a motile green flagellate, was collected off the north shore of New Guinea and maintained in culture in autoclaved seawater or an enriched seawater medium. In light, this green Noctiluca survived for at least a month and divided, without added food; while in darkness the symbiotic flagellate disappeared and the Noctiluca died within a few days, unless the food organism Dunaliella tertiolecta was present. Long-continued growth in light required the presence of Dunaliella, since the symbiotic flagellate was gradually lost under all the culture conditions assayed. All green Noctiluca assayed were luminescent. The light emitted per cell varied with the nutritional state of the host, and was as bright as a Japanese strain without symbionts when the Noctiluca was feeding on Dunaliella. In the absence of Dunaliella, the light emitted per Noctiluca depended on the light intensity in which cultures were maintained. The luminescent flash followed the same time course as that of other strains of Noctiluca without symbionts. No circadian rhythm of luminescence was present. Observations on both the growth and luminescence show that the flagellate symbiont contributes to the nutrition of the green Noctiluca. The symbiosis, however, is unstable in culture.     

Genetic effects of habitat fragmentation on blue sucker populations in the upper Missouri River (<Emphasis Type="Italic">Cycleptus elongatus</Emphasis> Lesueur, 1918)

The blue sucker, Cycleptus elongatus, is a large catostomid fish that occurs in main stem rivers throughout the Mississippi basin of North America. Although not federally listed as threatened or endangered, populations are not considered stable in any of 21 states where they occur. Included in the range is the Missouri River, which flows more than 3,200 km from Montana to St. Louis, Missouri. Historically, C. elongatus was distributed continuously throughout the main stem Missouri and its major tributaries, but from 1952 to 1963, six major impoundments were constructed on the upper Missouri by the US Army Corps of Engineers. The resulting reservoirs have inundated and fragmented large riverine habitat from Yankton, South Dakota to the headwaters. C. elongatus still occurs in remnant stretches between reservoirs; however, little is known of the impacts of the dams on these populations. In order to test for such effects, 231 individuals from nine sites were genotyped at 14 variable microsatellite loci. An additional 142 individuals from six sites in the Mississippi River were also genotyped for comparative purposes. In the Missouri, allelic richness was reduced in inter-reservoir sites relative to those in the free flowing lower river. In addition, significant isolation by distance occurs in the Missouri, a pattern not present in the unimpounded Mississippi. These results are consistent with reduced intradrainage gene flow in the Missouri River and are the first to indicate effects of impoundments on genetic structure in the system. This information will assist governing agencies in making informed decisions regarding conservation of C. elongatus in the Missouri River drainage and throughout the range.     

A novel microbial sensor using luminous bacteria.

A novel microbial sensor system that uses luminous bacteria was developed for the determination of both glucose and toxic compounds. The sensor system consisted of a membrane with luminous bacteria immobilized upon it and a photomultiplier. Measurements were based on the in vivo intensity of the light emitted by the bacteria, as this is affected by their environment. A linear relationship was observed between increased luminescence and concentrations of glucose between 0.05 mM and 0.55 mM. The relative standard deviation was 10% for 0.55 mM glucose (n = 10). Toxic compounds such as benzalkonium chloride, sodium dodecyl sulphate and chromium(VI) were also detected by measuring the decrease in luminescence in their presence.     

Functional morphology of the luminescence system of Siphamia versicolor (Perciformes: Apogonidae), a bacterially luminous coral reef fish

Previous studies of the luminescence system of Siphamia versicolor (Perciformes: Apogonidae) identified a ventral light organ, reflector, lens, duct, and a ventral diffuser extending from the throat to the caudal peduncle. The control and function of luminescence in this and other species of Siphamia, however, have not been defined. Morphological examination of fresh and preserved specimens identified additional components of the luminescence system involved in control and ventral emission of luminescence, including a retractable shutter over the ventral face of the light organ, contiguity of the ventral diffuser from the caudal peduncle to near the chin, and transparency of the bones and other tissues of the lower jaw. The shutter halves retract laterally, allowing the ventral release of light, and relax medially, blocking ventral light emission; topical application of norepinephrine to the exposed light organ resulted in retraction of the shutter halves, which suggests that operation of the shutter is under neuromuscular control. The extension of the diffuser to near the chin and transparency of the lower jaw allow a uniform emission of luminescence over the entire ventrum of the fish. The live aquarium‐held fish were found to readily and consistently display ventral luminescence. At twilight, the fish left the protective association with their longspine sea urchin, Diadema setosum, and began to emit ventral luminescence and to feed on zooplankton. Ventral luminescence illuminated a zone below and around the fish, which typically swam close to the substrate. Shortly after complete darkness, the fish stopped feeding and emitting luminescence. These observations suggest that S. versicolor uses ventral luminescence to attract and feed on zooplankton from the reef benthos at twilight. Ventral luminescence may allow S. versicolor to exploit for feeding the gap at twilight in the presence of potential predators as the reef transitions from diurnally active to nocturnally active organisms. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc     

Taxonomy of the marine,luminous bacteria

Summary One hundred and seventy-three strains of marine, luminous bacteria isolated from sea water, surfaces and intestines of fish, as well as from the luminous organs of fish and squid were submitted to an extensive phenotypic characterization. A numerical analysis of the results grouped these strains into four clusters which were formed on the basis of overall phenotypic similarity. One cluster, which was given the designationBeneckea harveyi, consisted of strains which had a moles% GC content in their DNAs of 46.5±1.3 and a single, sheathed, polar flagellum when grown in liquid medium. Most of these strains had unsheathed, peritrichous flagella in addition to the sheathed, polar flagellum when grown on solid medium. The two phenotypically similar clusters which were assigned the species designationsPhotobacterium phosphoreum andP. mandapamensis consisted of strains which had 1–3 unsheathed, polar flagella and moles % GC contents in their DNAs of 41.5±0.7 and 42.9±0.5, respectively. The cluster designatedP. fischeri contained strains having 2–8 sheathed, polar flagella and a moles % GC content of 39.8±1.1. These four species could be further distinguished on the basis of a number of nutritional properties as well as other phenotypic traits. The assignment of the luminous, marine bacteria to four species was supported by differences in the properties of the luminous system as well as differences in the pattern of regulation of spartokinase activity which are discussed. The speciesB. harveyi was found to be phenotypically similar to a number of previously characterized, non-luminous strains ofBeneckea which should probably be assigned to this species.Non-Standard Abbreviations ASW artificial sea water - ATCC American Type Culture Collection - BM basal medium - BMA basal medium agar - GC guanine plus cytosine - LA luminous medium agar - LB luminous medium broth - MA Difco Marine Agar - NCMB National Collection of Marine Bacteria - PHB poly--hydroxybutyrate - S similarity coefficient - YEB yeast extract broth This paper is part of a dissertation submitted by the senior author to the Graduate Division of the University of Hawaii in partial fulfillment of the requirements for the Ph.D. Degree in Microbiology     

STUDIES ON BIOLUMINESCENCE : XVII. FLUORESCENCE AND INHIBITION OF LUMINESCENCE IN CTENOPHORES BY ULTRA-VIOLET LIGHT.

1. Small dumps of the luminous cells of Mnemiopsis cannot readily be stimulated mechanically but will luminesce on treatment with saponin solution. Larger groups of luminous cells (such as are connected with two paddle plates) luminesce on mechanical stimulation. This suggests that mechanical stimulation to luminesce occurs chiefly through a nerve mechanism which has been broken up in the small dumps of luminous tissue. 2. The smallest bits of luminous tissue, even cells freed from the animal by agitation, that will pass through filter paper, lose their power to luminesce in daylight and regain it (at least partially) in the dark. 3. Luminescence of the whole animal and of individual cells is suppressed by near ultra-violet light (without visible light). 4. Inhibition in ultra-violet light is not due to stimulation (by the ultra-violet light) of the animal to luminesce, thereby using up the store of photogenic material. 5. Animals stimulated mechanically several times and placed in ultra-violet light show a luminescence along the meridians in the same positions as the luminescence that appears on stimulation. This luminescence in the ultra-violet or "tonic luminescence," is not obtained with light adapted ctenophores and is interpreted to be a fluorescence of the product of oxidation of the photogenic material. 6. Marked fluorescence of the luminous organ of the glowworm (Photuris) and of the luminous slime of Chatopterus may be observed in ultra-violet but no marked fluorescence of the luminous substances of Cypridina is apparent. 7. Evidence is accumulating to show a close relation between fluorescent and chemiluminescent substances in animals, similar to that described for unsaturated silicon compounds and the Grignard reagents.     

THE TOTAL LUMINOUS EFFICIENCY OF LUMINOUS BACTERIA

Methods are described for measuring the light emitted by an emulsion of luminous bacteria of given thickness, and calculating the light emitted by a single bacterium, measuring 1.1 x 2.2 micra, provided there is no absorption of light in the emulsion. At the same time, the oxygen consumed by a single bacterium was measured by recording the time for the bacteria to use up .9 of the oxygen dissolved in sea water from air (20 per cent oxygen). The luminescence intensity does not diminish until the oxygen concentration falls below 2 per cent, when the luminescence diminishes rapidly. Above 2 per cent oxygen (when the oxygen dissolving in sea water from pure oxygen at 760 mm. Hg pressure = 100 per cent) the bacteria use equal amounts of oxygen in equal times, while below 2 per cent oxygen it seems very likely that rate of oxygen absorption is proportional to oxygen concentration. By measuring the time for a tube of luminous bacteria of known concentration saturated with air (20 per cent oxygen) to begin to darken (2 per cent oxygen) we can calculate the oxygen absorbed by one bacterium per second. The bacteria per cc. are counted on a blood counting slide or by a centrifugal method, after measuring the volume of a single bacterium (1.695 x 10^–12 cc.). Both methods gave results in good agreement with each other. The maximum value for the light from a single bacterium was 24 x 10^–14 lumens or 1.9 x 10^–14 candles. The maximum value for lumen-seconds per mg. of oxygen absorbed was 14. The average value for lumen-seconds per mg. O₂ was 9.25. The maximum values were selected in calculating the efficiency of light production, since some of the bacteria counted may not be producing light, although they may still be using oxygen. The "diet" of the bacteria was 60 per cent glycerol and 40 per cent peptone. To oxidize this mixture each mg. of oxygen would yield 3.38 gm. calories or 14.1 watts per second. 1 lumen per watt is therefore produced by a normal bacterium which emits 14 lumen-seconds per mg. O₂ absorbed. Since the maximum lumens per watt are 640, representing 100 per cent efficiency, the total luminous efficiency if .00156. As some of the oxygen is used in respiratory oxidation which may have nothing to do with luminescence, the luminescence efficiency must be higher than 1 lumen per watt. Experiments with KCN show that this substance may reduce the oxygen consumption to 1/20 of its former value while reducing the luminescence intensity only ¼. A partial separation of respiratory from luminescence oxidations is therefore effected by KCN, and our efficiency becomes 5 lumens per watt, or .0078. This is an over-all efficiency, based on the energy value of the "fuel" of the bacteria, regarded as a power plant for producing light. It compares very favorably with the 1.6 lumens per watt of a tungsten vacuum lamp or the 3.9 lumens per watt of a tungsten nitrogen lamp, if we correct the usual values for these illuminants, based on watts at the lamp terminals, for a 20 per cent efficiency of the power plant converting the energy of coal fuel into electric current. The specific luminous emission of the bacteria is 3.14 x 10^–6 lumens per cm². One bacterium absorbs 215,000 molecules of oxygen per second and emits 1,280 quanta of light at λ_max = 510µµ. If we suppose that a molecule of oxygen uniting with luminous material gives rise to the emission of 1 quantum of light energy, only 1/168 of the oxygen absorbed is used in luminescence. On this basis the efficiency becomes 168 lumens per watt or 26.2 per cent.     

Research into bioluminescence of the Black Sea ctenophores Pleurobrachia pileus O.F. Müller, 1776

The work presents the findings of the laboratory and in situ studies of ctenophore Pleurobrachia pileus O.F. Müller, 1776 which have shown that this species has bioluminescent properties. These organisms were considered non-luminous before. P. pileus bioluminescence was studied on board RV Professor Vodyanitsky during its 116th voyage. Sampling live organisms was preceded by probing with the Salpa MA+ probe to register the daily maximum glow in redoxcline, which in this zone was recorded, as a rule, in the depth range of 60–70 m, where dense clusters of P. pileus were formed at that time. The samples of ctenophores were taken by a Bogorov–Rass plankton net. After the net was closed, it was lifted to the surface at a speed of 0.4–0.5 m s⁻¹. It was shown that only at a temperature not exceeding 14°C, the P. pileus remained alive for 2–3 days. The data provided indicate that the temperature above 14°C is close to the maximum permissible for P. pileus; therefore, chemical and mechanical stimulation experiments were carried out at this temperature (14°C) to agitate ctenophores luminescence. Though, the nature of their signal was significantly different. The total percentage of luminous organisms from the entire catch was 32.43%, which unequivocally proves that P. pileus glows and makes a significant contribution to the intensity of the glow at great depths in redoxcline.     

Phylogenetic analysis of host–symbiont specificity and codivergence in bioluminescent symbioses

Several groups of marine fishes and squids form mutualistic bioluminescent symbioses with luminous bacteria. The dependence of the animal on its symbiont for light production, the animal's specialized anatomical adaptations for harboring bacteria and controlling light emission, and the host family bacterial species specificity characteristic of these associations suggest that bioluminescent symbioses are tightly coupled associations that might involve coevolutionary interactions. Consistent with this possibility, evidence of parallel cladogenesis has been reported for squid–bacterial associations. However, genetic adaptations in the bacteria necessary for and specific to symbiosis have not been identified, and unlike obligate endosymbiotic associations in which the bacteria are transferred vertically, bacterially bioluminescent hosts acquire their light‐organ symbionts from the environment with each new host generation. These contrasting observations led us to test the hypotheses of species specificity and codivergence in bioluminescent symbioses, using an extensive sampling of naturally formed associations. Thirty‐five species of fish in seven teleost families (Chlorophthalmidae, Macrouridae, Moridae, Trachichthyidae, Monocentridae, Acropomatidae, Leiognathidae) and their light‐organ bacteria were examined. Phylogenetic analysis of a taxonomically broad sampling of associations was based on mitochondrial 16S rRNA and cytochrome oxidase I gene sequences for the fish and on recA, gyrB and luxA sequences for bacteria isolated from the light organs of these specimens. In a fine‐scale test focused on Leiognathidae, phylogenetic analysis was based also on histone H3 subunit and 28S rRNA gene sequences for the fish and on gyrB, luxA, luxB, luxF and luxE sequences for the bacteria. Deep divergences were revealed among the fishes, and clear resolution was obtained between clades of the bacteria. In several associations, bacterial species identities contradicted strict host family bacterial species specificity. Furthermore, the fish and bacterial phylogenies exhibited no meaningful topological congruence; evolutionary divergence of host fishes was not matched by a similar pattern of diversification in the symbiotic bacteria. Re‐analysis of data reported for squids and their luminous bacteria also revealed no convincing evidence of codivergence. These results refute the hypothesis of strict host family bacterial species specificity and the hypothesis of codivergence in bioluminescent symbioses. © The Willi Hennig Society 2007.