Glowing in the dark: discriminating patterns of bioluminescence from different taxa during the Arctic polar night

Research since 2009 has shown that despite almost total darkness during the Arctic polar night, there is much more biological activity than previously assumed, both at the sea surface, water column and sea floor. Here, we describe in situ monitoring of the bioluminescent fraction of the zooplankton community (dinoflagellates, copepods, krill and ctenophores) as a function of time and space. In order to examine the relative contribution of each selected taxon and any diurnal patterns in the relative signals, a time series platform capable of detecting in situ bioluminescent flashes was established in Kongsfjord, Svalbard, during the polar night in January 2013. Combined with laboratory-controlled measurements of animals collected next to the time series platform, we present both taxon-specific and community characteristics of the bioluminescence signal from a location at 79°N and from the middle of the polar night. Based on this 51-h time series, we conclude that the bioluminescent fraction of the zooplankton does not maintain a diurnal signal. Rather, the frequency of bioluminescence flashes from the entire bioluminescent community remained steady throughout the sampling period. Furthermore, we conclude that bioluminescence flash kinetic characteristics have a strong potential for in situ taxa recognition of zooplankton.     

Fading bioluminescence of the tropical Atlantic Ocean

Bioluminescence intensity acts as the indicator of the functional state of a plankton community. Data on bioluminescence intensity, zooplankton biomass, and chlorophyll a from the expeditions to the tropical and subtropical Atlantic Ocean were analyzed. The regression models implied a ~10-fold decline of bioluminescence intensity [bioluminescence potential (BP)] and zooplankton biomass for the 46 year time range from 1970 to 2016. The correlation was low between chlorophyll a (the indicator of phytoplankton biomass) and BP, but it was significant for the zooplankton biomass BP annual time series. The decline of BP was associated with the decreasing abundance of bioluminescent zooplankton, and increasing global warming-driven temperature.     

Bioluminescent eddies of the World Ocean

Mesoscale eddies of the ocean (with a characteristic diameter of about 100 km and a life time-span of about several weeks) are habitats of plankton organisms, many of which are bioluminescent. The spatial heterogeneity of bioluminescence of the upper mixed layer associated with the impact of mesoscale eddies is poorly studied. The 45-year historical data set was retrieved, in order to select the bathy-photometric surveys carried out in the form of station grids and transects across eddies. Data from 71 expeditions deployed in 1966–2022 to the Atlantic Ocean, Indian Ocean and Mediterranean Sea basin were analyzed, in order for the spatial heterogeneity of bioluminescent fields to be elucidated across eddy fields. The stimulated bioluminescence intensity was characterized by the bioluminescent potential, which represented the maximal amount of radiant energy emitted in a given volume of water by bioluminescent organisms. The normalized bioluminescent potential over oceanographic station grids exhibited correlation with the eddy kinetic energy and zooplankton biomass (r = 0.8, at P = 0.001 and r = 0.7, at P = 0.05, respectively), in a broad range of energy and bioluminescence units (0.02–0.2 m² s⁻²; 0.4–92.0 × 10⁻⁸ W cm⁻² L⁻¹, respectively). Overall, estimates of bioluminescent potential variability on the mesoscale contribute to the assessment of the multiple-scale variation of the bioluminescent field of the World Ocean.     

MICROSOURCES OF BIOLUMINESCENCE IN PYROCYSTIS FUSIFORMIS (PYRROPHYTA)1

The large dinoflagellate, Pyrocystis fusiformis Murray, emits biolumtnescence on stimulation with dilute acid. The bioluminescence can be seen in the light microscope to originate in a spherical region just distal to the nucleus during the day and appears as a persistent glow which can be localized in an orange-brown sphere. At night, the bioluminescence, in response to stimulation, is a bright flash from microsources scattered throughout the cytoplasm. The orange sphere can no longer be seen nor does a bioluminescent glow originate from this central region on stimulation. This difference in the position of intracellular bioluminescence between day and night has allowed the identification in electron micrographs of structures which correspond to the source of bioluminescence during the day. Light is emitted from a spherical mass of vesicles which contain electron-dense short rods with rounded ends, sometimes crossed by electron-transparent narrow bands. At night, these vesicles can be recognized in the peripheral cytoplasm. It is proposed that these vesicles are the structural counterparts of the microsources of bioluminescence in P. fusiformis.     

Exogenous compounds in studying the mechanism of electron-excited state formation in bioluminescence

Using a series of exogenous fluorescent molecules as potential energy acceptors, the hypothesis on the activity of the upper electron-excited states in bioluminescence was tested. The results in bacterial and firefly bioluminescent enzyme systems were compared. Similar activity to the energetic precursor in bacterial bioluminescence was not proven in the case of the firefly system, the result of a very efficient intramolecular energy transfer in the emitter of the firefly bioluminescence. The influence of a number of metallic salts on a bacterial bioluminescent enzyme system was studied. Bioluminescence inhibition coefficients were compared to the free energies of electron withdrawing of cations. The correlation shows that inhibition and activation of luminescence intensity result from the effects of cations on electron transfer in the bioluminescent system.     

Use of bioluminescent Escherichia coli O157:H7 to study intra-protozoan survival of bacteria within Tetrahymena pyriformis

A method was developed that enabled real-time monitoring of the uptake and survival of bioluminescent Escherichia coli O157 within the freshwater ciliate Tetrahymena pyriformis. Constitutively bioluminescent E. coli O157 pLITE27 was cocultured with T. pyriformis in nutrient-deficient (Chalkley's) and in nutrient-rich (proteose peptone, yeast extract) media. Non-internalised bacteria were inactivated by addition of colistin, indicated by a decline in bioluminescence. Protozoa were subsequently lysed with Triton X-100 which lead to a further drop in bioluminescence, consistent with release of live internal bacteria from T. pyriformis into the colistin-containing environment. Bioluminescence measurements for non-lysed cultures indicated that internalised E. coli O157 pLITE27 cells were only slowly digested by T. pyriformis, in both media, over the time period studied. The results suggest that bioluminescent bacteria are useful tools in the study of bacterial intra-protozoan survival.     

Exploring the signaling pathway of circadian bioluminescence

Bioluminescence in the unicellular dinoflagellate Gonyaulax polyedra represents an excellent model for studying a circadian controlled process at the biochemical and molecular levels. There are three key components involved in the bioluminescence reaction: the enzyme, luciferase, its substrate, luciferin, and a luciferin-binding protein (LBP), which sequesters the substrate at p^H 7.5 and thus prevents it from reacting with the enzyme. All components are tightly packed together in organdies, designated scintillons. The entire bioluminescent system is under circadian control with maximum amounts in the night. For both proteins circadian control is exerted at the translational level. In case of Ibp mRNA a small interval in its 3'untranslated region serves as a cis -acting element to which a trans -factor binds in a circadian manner. The binding activity of this factor decreases at the beginning of the night phase, when synthesis of LBP starts, and it increases al the end of the night, when synthesis of LBP stops indicating that it functions as a clock-controlled represser.     

萤火虫(鞘翅目:萤科)两性交流中的闪光信号

对国内外萤火虫两性交流闪光信号的研究进行了综述,萤火虫发光器因种而异,多数发出黄绿色萤光,闪光信号的频率、光谱、强度及其时空分布的闪光模式包含着两性交流信息。萤火虫闪光交流系统有两种分类方法,其一是萤火虫具两个类型的闪光信号交流系统,及系统和系统,前者多在旧大陆,后者多在新大陆;其二是萤火虫具6个类型闪光信号交流系统,即HP,LL,LC,PR,CR和LB型,其中PR型与系统相对应,HP型与系统对应。萤火虫两性交流闪光信号常因时间和空间上的差异及外界物体的干扰使两性闪光交流的效率受到影响。萤火虫两性交流的闪光信号起源于鞘翅目的幼虫阶段,并起警戒天敌的作用,经过两性选择成为成虫两性交流的一种途径,进而成为新大陆的一些萤火虫间捕食猎物和逃避天敌的生存策略。     

Phylogenomic relationships of bioluminescent elateroids define the ‘lampyroid’ clade with clicking Sinopyrophoridae as its earliest member

Bioluminescence has been hypothesized as aposematic signalling, intersexual communication and a predatory strategy, but origins and relationships among bioluminescent beetles have been contentious. We reconstruct the phylogeny of the bioluminescent elateroid beetles (i.e. Elateridae, Lampyridae, Phengodidae and Rhagophthalmidae), analysing genomic data of Sinopyrophorus Bi & Li, and in light of our phylogenetic results, we erect Sinopyrophoridae Bi & Li, stat.n . as a clicking elaterid‐like sister group of the soft‐bodied bioluminescent elateroid beetles, that is, Lampyridae, Phengodidae and Rhagophthalmidae. We suggest a single origin of bioluminescence for these four families, designated as the ‘lampyroid clade’, and examine the origins of bioluminescence in the terminal lineages of click beetles (Elateridae). The soft‐bodied bioluminescent lineages originated from the fully sclerotized elateroids as a derived clade with clicking Sinopyrophorus and Elateridae as their serial sister groups. This relationship indicates that the bioluminescent soft‐bodied elateroids are modified click beetles. We assume that bioluminescence was not present in the most recent common ancestor of Elateridae and the lampyroid clade and it evolved among this group with some delay, at the latest in the mid‐Cretaceous period, presumably in eastern Laurasia. The delimitation and internal structure of the elaterid‐lampyroid clade provides a phylogenetic framework for further studies on the genomic variation underlying the evolution of bioluminescence.     

Characterization of bioluminescent derivatives of assimilable organic carbon test bacteria

The assimilable organic carbon (AOC) test is a standardized measure of the bacterial growth potential of treated water. We describe the design and initial development of an AOC assay that uses bioluminescent derivatives of AOC test bacteria. Our assay is based on the observation that bioluminescence peaks at full cell yield just prior to the onset of the stationary phase during growth in a water sample. Pseudomonas fluorescens P-17 and Spirillum sp. strain NOX bacteria were mutagenized with luxCDABE operon fusion and inducible transposons and were selected on minimal medium. Independent mutants were screened for high luminescence activity and predicted AOC assay sensitivity. All mutants tested were able to grow in tap water under AOC assay conditions. Strains P-17 I5 (with p-aminosalicylate inducer) and NOX I3 were chosen for use in the bioluminescence AOC test. Peak bioluminescence and plate count AOC were linearly related for both test bacteria, though data suggest that the P-17 bioluminescence assay requires more consistent luminescence monitoring. Bioluminescence results were obtained 2 or 3 days postinoculation, compared with 5 days for the ATP luminescence AOC assay and 8 days for the plate count assay. Plate count AOC assay results for nonmutant and bioluminescent bacteria from 36 water samples showed insignificant differences, indicating that the luminescent bacteria retained a full range of AOC measurement capability. This bioluminescence method is amenable to automation with a microplate format with programmable reagent injection.     

Study of the binding between bacterial lipopolysaccharides and polymyxin by a bioluminescent method

For rating the interaction of lipopolysaccharides (LPS) with polymixin B (PmB) a bacterial bioluminescence is offered to be used. Bioluminescence level of bacteria decreases under free antibiotic amounts action. It is shown, that as a result of interaction with LPS antibiotic properties of PmB are reduced, and the intensity of bacterial bioluminescence is restored. The bioluminescence level in such system characterizes the LPS quantity. Kinetic properties of bacterial light emission at the presence of LPS and PmB are investigated as well as equilibrium state of the system. Kinetic and equilibrium constants describing this reaction are determined. The conditions of quantitative bioluminescent definition of LPS in an interval of concentration 0.166-10 micrograms/ml have been chosen and calibration curves are presented.     

In vivo bioluminescence imaging

In vivo bioluminescent imaging (BLI) is a versatile and sensitive tool that is based on detection of light emission from cells or tissues. Bioluminescence, the biochemical generation of light by a living organism, is a naturally occurring phenomenon. Luciferase enzymes, such as that from the North American firefly (Photinus pyralis), catalyze the oxidation of a substrate (luciferin), and photons of light are a product of the reaction. Optical imaging by bioluminescence allows a low-cost, noninvasive, and real-time analysis of disease processes at the molecular level in living organisms. Bioluminescence has been used to track tumor cells, bacterial and viral infections, gene expression, and treatment response. Bioluminescence in vivo imaging allows longitudinal monitoring of a disease course in the same animal, a desirable alternative to analyzing a number of animals at many time points during the course of the disease. We provide a brief introduction to BLI technology, specific examples of in vivo BLI studies investigating bacterial/viral pathogenesis and tumor growth in animal models, and highlight some future perspectives of BLI as a molecular imaging tool.     

A Causal Relation between Bioluminescence and Oxygen to Quantify the Cell Niche

Bioluminescence imaging assays have become a widely integrated technique to quantify effectiveness of cell-based therapies by monitoring fate and survival of transplanted cells. To date these assays are still largely qualitative and often erroneous due to the complexity and dynamics of local micro-environments (niches) in which the cells reside. Here, we report, using a combined experimental and computational approach, on oxygen that besides being a critical niche component responsible for cellular energy metabolism and cell-fate commitment, also serves a primary role in regulating bioluminescent light kinetics. We demonstrate the potential of an oxygen dependent Michaelis-Menten relation in quantifying intrinsic bioluminescence intensities by resolving cell-associated oxygen gradients from bioluminescent light that is emitted from three-dimensional (3D) cell-seeded hydrogels. Furthermore, the experimental and computational data indicate a strong causal relation of oxygen concentration with emitted bioluminescence intensities. Altogether our approach demonstrates the importance of oxygen to evolve towards quantitative bioluminescence and holds great potential for future microscale measurement of oxygen tension in an easily accessible manner.     

Real-time compensation of the inner filter effect in high-density bioluminescent cultures

Bioluminescence has recently become a popular research tool in several fields, including medicine, pharmacology, biochemistry, bioprocessing, and environmental engineering. Beginning with purely qualitative goals, scientists are now targeting more demanding applications where accurate, quantitative interpretation of bioluminescence is necessary. Using the recent advances in fiber-optic technology, bioluminescence is easily monitored in vivo and in real time. However, the convenience of this measurement is often concealing an unsuspected problem: the bioluminescence signal might be corrupted by a large error caused by the extinction of light by biological cells. Since bioluminescent cultures not only emit light but also absorb and scatter it, the measured signal is related in a complex, nonlinear, and cell-concentration-dependent manner to the "true" bioluminescence. This light extinction effect, known as the "inner filter effect," is significant in high-density cultures. Adequate interpretation of the bioluminescence signal can be difficult without its correction. Here, we propose a real-time algorithm for elimination of the inner filter effect in a bioreactor. The algorithm yields the bioluminescence which would be measured if the glowing culture was completely transparent. This technique has been successfully applied to batch and continuous cultivation of recombinant bioluminescent Escherichia coli. (c) 1993 John Wiley & Sons, Inc.     

CCD-monitoring of bioluminescence during the induction of the cell wall-deficient, L-form state of a genetically modified strain of Pseudomonas syringae pv. phaseolicola

Bioluminescence from developing L-form colonies of the plant pathogen, Pseudomonas syringae pv. phaseolicola , was monitored using the enhanced light-detecting capabilities of a charge-coupled device. During L-form induction, the bacteria entered a prolonged period during which the level of light output and hence metabolic activity, was very low. A relatively small number of highly bioluminescent L-form colonies were then observed to develop against a background of non-bioluminescent bacteria. When these colonies were sub-cultured and examined microscopically, typical L-form morphology was observed and continued high bioluminescence was detectable from derived colonies.     

Circadian expression of the dnaK gene in the cyanobacterium Synechocystis sp. strain PCC 6803.

The expression of the dnaK gene in the cyanobacterium Synechocystis sp. strain PCC 6803 was continuously monitored as bioluminescence by an automated monitoring system, using the bacterial luciferase genes (luxAB) of Vibrio harveyi as a reporter of promoter activity. A dnaK-reporting bioluminescent Synechocystis strain was constructed by fusing a promoterless segment of the luxAB gene set downstream of the promoter region of the Synechocystis dnaK gene and introduction of this gene fusion into a BglII site downstream of the ndhB gene in the Synechocystis chromosome. Bioluminescence from this strain was continuously monitored and oscillated with a period of about 22 h for at least 5 days in continuous light. The phase of the rhythm was reset by the timing of the 12-h dark period administered prior to the continuous light. The period of the rhythm was temperature compensated between 25 and 35 degrees C. Thus, the bioluminescence rhythm satisfied the three criteria of circadian rhythms. Furthermore, the abundance of dnaK mRNA also oscillated with a period of about 1 day for at least 2 days in continuous light conditions, indicating circadian control of dnaK gene expression in Synechocystis sp. strain PCC 6803.     

Bioluminescent organs of two deep-sea arrow worms, Eukrohnia fowleri and Caecosagitta macrocephala, with further observations on Bioluminescence in chaetognaths

Bioluminescence in the deep-sea chaetognath Eukrohnia fowleri is reported for the first time, and behavioral, morphological, and chemical characteristics of bioluminescence in chaetognaths are examined. Until this study, the only known species of bioluminescent chaetognath was Caecosagitta macrocephala. The luminescent organ of that species is located on the ventral edge of each anterior lateral fin, whereas that of E. fowleri runs across the center of the tail fin on both dorsal and ventral sides. Scanning electron microscopy showed that the bioluminescent organs of both species consist of hexagonal chambers containing elongate ovoid particles-the organelles holding bioluminescent materials. No other luminous organism is known to use hexagonal packing to hold bioluminescent materials. Transmission electron microscopy of particles from C. macrocephala revealed a densely packed paracrystalline matrix punctuated by globular inclusions, which likely correspond to luciferin and luciferase, respectively. Both species use unique luciferases in conjunction with coelenterazine for light emission. Luciferase of C. macrocephala becomes inactive after 30 min, but luciferase of E. fowleri is highly stable. Although C. macrocephala has about 90 times fewer particles than E. fowleri, it has a similar bioluminescent capacity (total particle volume) due to its larger particle size. In situ observations of C. macrocephala from a remotely operated vehicle revealed that the luminous particles are released to form a cloud. The discovery of bioluminescence in a second chaetognath phylogenetically distant from the first highlights the importance of bioluminescence among deep-sea organisms.     

Characterization of Bioluminescent Derivatives of Assimilable Organic Carbon Test Bacteria

The assimilable organic carbon (AOC) test is a standardized measure of the bacterial growth potential of treated water. We describe the design and initial development of an AOC assay that uses bioluminescent derivatives of AOC test bacteria. Our assay is based on the observation that bioluminescence peaks at full cell yield just prior to the onset of the stationary phase during growth in a water sample. Pseudomonas fluorescens P-17 and Spirillum sp. strain NOX bacteria were mutagenized with luxCDABE operon fusion and inducible transposons and were selected on minimal medium. Independent mutants were screened for high luminescence activity and predicted AOC assay sensitivity. All mutants tested were able to grow in tap water under AOC assay conditions. Strains P-17 I5 (with p-aminosalicylate inducer) and NOX I3 were chosen for use in the bioluminescence AOC test. Peak bioluminescence and plate count AOC were linearly related for both test bacteria, though data suggest that the P-17 bioluminescence assay requires more consistent luminescence monitoring. Bioluminescence results were obtained 2 or 3 days postinoculation, compared with 5 days for the ATP luminescence AOC assay and 8 days for the plate count assay. Plate count AOC assay results for nonmutant and bioluminescent bacteria from 36 water samples showed insignificant differences, indicating that the luminescent bacteria retained a full range of AOC measurement capability. This bioluminescence method is amenable to automation with a microplate format with programmable reagent injection.     

Bioluminescence Imaging Reveals Dynamics of Beta Cell Loss in the Non-Obese Diabetic (NOD) Mouse Model

We generated a mouse model (MIP-Luc-VU-NOD) that enables non-invasive bioluminescence imaging (BLI) of beta cell loss during the progression of autoimmune diabetes and determined the relationship between BLI and disease progression. MIP-Luc-VU-NOD mice displayed insulitis and a decline in bioluminescence with age which correlated with beta cell mass, plasma insulin, and pancreatic insulin content. Bioluminescence declined gradually in female MIP-Luc-VU-NOD mice, reaching less than 50% of the initial BLI at 10 weeks of age, whereas hyperglycemia did not ensue until mice were at least 16 weeks old. Mice that did not become diabetic maintained insulin secretion and had less of a decline in bioluminescence than mice that became diabetic. Bioluminescence measurements predicted a decline in beta cell mass prior to the onset of hyperglycemia and tracked beta cell loss. This model should be useful for investigating the fundamental processes underlying autoimmune diabetes and developing new therapies targeting beta cell protection and regeneration.