Circadian clock period length is not consistently linked to chronotype in a wild songbird.

Circadian clock properties vary between individuals and relate to variation in entrained timing in captivity. How this variation translates into behavioural differences in natural settings, however, is poorly understood. Here, we tested in great tits whether variation in the free-running period length (tau) under constant dim light (LL) was linked to the phase angle of the entrained rhythm ("chronotype") in captivity and in the wild, as recently indicated in our study species. We also assessed links between tau and the timing of first activity onset and offset under LL relative to the last experienced light-dark (LD) cycle. We kept 66 great tits, caught in two winters, in LL for 14 days and subsequently released them with a radio transmitter back to the wild, where their activity and body temperature rhythms were tracked for 1 to 22 days. For a subset of birds, chronotype was also recorded in the lab before release. Neither wild nor lab chronotypes were related to tau. We also found no correlation between lab and wild chronotypes. However, the first onset in LL had a positive relationship with tau, but only in males. Our results demonstrate that links between tau and phase of entrainment, postulated on theoretical grounds, may not consistently hold under natural conditions, possibly due to strong masking. This calls for more holistic research on how the many components of the circadian system interact with the environment to shape timing in the wild. Wild birds showed chronotypes in the field that were unlinked to their circadian period length tau measured in captivity. In males only, the first onset of activity after exposure to constant dim light did correlate with tau. Our study emphasises the need to investigate clocks in the real world, including a need to better understand masking.

Microbial nitrogen removal in reef-building corals: A light-sensitive process.

Scleractinian corals are the main framework-building groups in tropical coral reefs. In the coral holobiont, nitrogen-cycling mediated by microbes is fundamental for sustaining the coral reef ecosystems. However, little direct evidence characterizing the activities of microbial nitrogen removal via complete denitrification and anaerobic ammonium oxidation (anammox) in stony corals has been presented. In this study, multiple incubation experiments using 15N-tracer were conducted to identify and characterize N2 production by denitrification and anammox in the stony coral Pocillopora damicornis. The rates of denitrification and anammox were recorded up to 0.765 ± 0.162 and 0.078 ± 0.009 nmol N2 cm-2 h-1 respectively. Denitrification contributed the majority (∼90%) of N2 production by microbial nitrogen removal in stony corals. The microbial nitrogen removal activities showed diel rhythms, which might correspond to photosynthetic oxygen production. The N2 production rates of anammox and denitrification increased with incubation time. To the authors' knowledge, this study is the first to confirm and characterize the activities of complete denitrification and anammox in stony corals via stable isotope techniques. This study extends the understanding on nitrogen-cycling in coral reefs and how it participates in corals' resilience to environmental stressors.

Spontaneous recurrence of a summer-like diel rhythm in the body temperature of the Syrian hamster after hibernation.

Mammalian hibernation is a survival strategy characterized by metabolic suppression and drastically lowering body temperature (Tb), used during harsh seasons with food shortages and cold. The Syrian hamster commences hibernation in response to a short photoperiod and cold but spontaneously concludes hibernation after several months without environmental cues. Little is known about the changes in diel rhythms during hibernation. Using long-term and high-resolution Tb data, we analysed the diel Tb rhythm time-course changes in Syrian hamsters raised under summer-like conditions (long photoperiod (LP) and warm; LP-warm) and transferred to winter-like conditions (short photoperiod (SP) and cold; SP-cold). The diel Tb rhythm was undetectable during the hibernation period (HIBP), reappearing after the HIBP. The phase of this returning rhythm reverted to the LP entrainment phase characteristics despite the ambient SP and then re-entrained to the ambient SP as if the hamsters were transferred from the LP-warm to SP-cold conditions. The diel Tb rhythm reverted from the SP- to LP-type in a hibernation-dependent manner. Under constant dark and cold conditions, the circadian Tb rhythm recovered without photic stimuli following the HIBP. These findings suggest that hibernation involves a program that anticipates the ambient photoperiod when animals emerge from hibernation.

'Phototaxis' in the absence of light? Locomotory patterns in unionid mussels.

Locomotion in benthic invertebrates can strongly affect habitat selection and ecosystem nutrient cycling. In the case of freshwater mussels, the drivers of locomotion are largely unresolved. Our aim was to assess the influence of light presence and intensity on the locomotory behaviour of freshwater mussels in controlled laboratory experiments. The species investigated in our study were Anodonta anatina and Unio pictorum, two widely distributed mussels in European lentic and lotic inland waters. At low algal concentrations, known to be associated with more frequent locomotory activities, we found that both species moved primarily in the absence of light (72.7% of all movements across experiments). However, the movements of both species were directed towards the light source, resembling a net-positive 'phototactic' response but in the absence of light. The distance to the light source, which was negatively correlated to light intensity, had a positive effect on the distance covered in locomotory activities by A. anatina but not by U. pictorum. Intraspecific variation in shell size had no impact on movement distance, indicating that the energetic costs of movement were not a limiting factor. We suggest that the observed movement towards brighter locations helps to enhance food quantity and quality, whilst movement in darkness mitigates predation risks.

Diurnal gene oscillations modulated by RNA metabolism in tomato.

Diurnal rhythms are known to regulate the expression of a large number of genes, coordinating plant growth and development with diel changes in light and temperature. However, the impact of RNA metabolism on rhythmic gene oscillations in plant is not yet fully understood. To address this question, we performed transcriptome and degradome profiling on tomato leaves at 6 time points during one 24 h cycle, using RNA-seq and genome-wide mapping of uncapped and cleavage transcripts (GMUCT). Time-series profiling of RNA-seq revealed 9342 diurnal-oscillated genes, which were enriched in various metabolic processes. To quantify the general level of RNA degradation for each gene, we utilized the Proportion Uncapped (PU) metric, which represents the GMUCT/RNA-seq ratio. Oscillated PU analysis revealed that 3885 genes were regulated by rhythmic RNA degradation. The RNA decay of these diurnal genes was highly coordinated with mRNA downregulation during oscillation, highlighting the critical role of internal transcription-degradation balance in rhythmic gene oscillation. Furthermore, we identified 2190 genes undergoing co-translational RNA decay (CTRD) with 5' phosphate read ends enriched at the boundary of ribosomes stalling at translational termination sites. Interestingly, diurnal-changed mRNAs with large amplitudes tended to be co-translationally decay, suggesting that CTRD contributed to the rapid turnover of diurnal mRNAs. Finally, we also identified several genes, whose miRNA cleavage efficiency oscillated in a diurnal manner. Taken together, these findings uncovered the vital functions of RNA metabolism, including rhythmic RNA degradation, CTRD, and miRNA cleavage, in modulating the diurnal mRNA oscillations during diel change at post-transcriptional level in tomato.

Research on Photoinduction-Based Technology for Trapping Asian Longhorned Beetle (Anoplophora glabripennis (Motschulsky, 1853) (Coleoptera: Cerambycidae).

Light traps play a crucial role in monitoring pest populations. However, the phototactic behavior of adult Asian longhorned beetle (ALB) remains enigmatic. To provide a theoretical foundation to select the suitable light emitting diode (LED)-based light sources used for monitoring ALB, we compared the effect of exposure time on the phototactic response rates of adults at wavelengths of 365 nm, 420 nm, 435 nm, and 515 nm, and found that the phototactic rate increased gradually when the exposure time was prolonged, but there was no significant difference between different exposure times. We evaluated the effect of diel rhythm and found the highest phototactic rate at night (0:00-2:00) under 420 nm and 435 nm illumination (74-82%). Finally, we determined the phototactic behavioral response of adults to 14 different wavelengths and found both females and males showed a preference for violet wavelengths (420 nm and 435 nm). Furthermore, the effect of the light intensity experiments showed that there were no significant differences in the trapping rate between different light intensities at 120 min exposure time. Our findings demonstrate that ALB is a positively phototactic insect, showing that 420 nm and 435 nm are the most suitable wavelengths for attracting adults.

Butterfly foraging is remarkably synchronous in an experimental tropical macrocosm.

Diel patterns in foraging activity are dictated by a combination of abiotic, biotic and endogenous limits. Understanding these limits is important for insects because ectotherm taxa will respond more pronouncedly to ongoing climatic change, potentially affecting crucial ecosystem services. We leverage an experimental macrocosm, the Montreal Insectarium Grand Vivarium, to test the importance of endogenous mechanisms in determining temporal patterns in foraging activity of butterflies. Specifically, we assessed the degree of temporal niche partitioning among 24 butterfly species originating from the Earth's tropics within controlled environmental conditions. We found strong niche overlap, with the frequency of foraging events peaking around solar noon for 96% of the species assessed. Our models suggest that this result was not due to the extent of cloud cover, which affects radiational heating and thus limits body temperature in butterflies. Together, these findings suggest that an endogenous mechanism evolved to regulate the timing of butterfly foraging activity within suitable environmental conditions. Understanding similar mechanisms will be crucial to forecast the effects of climate change on insects, and thus on the many ecosystem services they provide.

Life history strategy dictates thermal preferences across the diel cycle and in response to starvation in variable field crickets, Gryllus lineaticeps.

Insects behaviorally thermoregulate across the diel cycle, and their preferred microhabitats change based on current resources available and the thermal performance optima of traits. Specific combinations of traits being prioritized are set by life history strategies, making life history an important intrinsic determinant of thermal preferences. However, we do not know how life history strategies shape plasticity of behavioral thermoregulation, limiting our ability to predict responses to environmental variability. We compared female variable field crickets (Gryllus lineaticeps) that are flight-capable (long-winged) and flightless (short-winged) to test the hypothesis that life history strategy determines plasticity of thermal preferences across the diel cycle and following starvation. Thermal preferences were elevated during the nocturnal activity period, and long-winged crickets preferred warmer temperatures compared to short-winged crickets across the diel cycle when fully fed. However, thermal preferences of starved crickets were reduced compared to fed crickets. The reduction in thermal preferences was greater in long-winged crickets, resulting in similar thermal preferences between starved long- and short-winged individuals and reflecting a more plastic response. Thus, life history does determine plasticity in thermoregulatory behaviors following resource limitations and effects of life history on thermal preferences are context dependent.

Diel and monthly activity pattern of brown bears and sika deer in the Shiretoko Peninsula, Hokkaido, Japan.

Mammals exhibit several types of diel activity pattern, including nocturnal, diurnal, crepuscular, and cathemeral. These patterns vary inter- and intra-specifically and are affected by environmental factors, individual status, and interactions with other individuals or species. Determining the factors that shape diel activity patterns is challenging but essential for understanding the behavioral ecology of animal species, and for wildlife conservation and management. Using camera-trap surveys, we investigated the species distributions and activity patterns of terrestrial mammals on the Shiretoko Peninsula, Hokkaido, Japan, with particular focus on brown bears and sika deer. From June to October 2019, a total of 7,530 observations were recorded by 65 camera-traps for eight species, including two alien species. The diel activity pattern of brown bears was diurnal/crepuscular, similar to that of bears in North America, but different from European populations. Bear observations were more frequent during the autumnal hyperphagia period, and adult females and sub-adults were more diurnal than adult males. In addition, bears inside the protected area were more diurnal than those outside it. These findings suggest that appetite motivation, competitive interactions between conspecifics, and human activities potentially affect bear activity patterns. Similar to other sika deer populations and other deer species, the diel activity patterns of sika deer were crepuscular. Deer showed less variation in activity patterns among months and sex-age classes, while adult males were observed more frequently during the autumn copulation period, suggesting that reproductive motivation affects their activity patterns.

Detailed Analysis of Zebrafish Larval Behaviour in the Light Dark Challenge Assay Shows That Diel Hatching Time Determines Individual Variation.

Research on stress coping style, i.e., the response of an organism to adverse conditions, which is constant over time and context, gained momentum in recent years, to better understand behavioural patterns in animal welfare. However, knowledge about the ontogeny of stress coping style is still limited. Here, we performed a detailed analysis of the light dark challenge behavioural assay in zebrafish larvae, where after acclimation in ambient light sudden alternating dark and light phases elicit an anxiety-like response. A principal component analysis on parameters related to locomotion (distance moved, swimming velocity, acceleration, mobility) and directionality (angular velocity, meandering of swimming path) revealed independence between the parameters determined in the light and the dark phases of the assay, indicating unrelated generalised behaviours per phase. However, high collinearity was observed between behavioural parameters within the same phase, indicating a robust response to the stimulus within behavioural phenotypes. Subsequently, this assay was used to determine the correlation between individual hatching time and the behavioural phenotype. The results show that fish that had hatched during daytime have a stronger behavioural response to the dark phase at 5 days post-fertilisation in locomotion related parameters and a weaker response in directionality related parameters, than fish that had hatched during nighttime. These results show that behavioural responses to the light dark challenge assay are robust and can be generalised for the light and the dark phase, and that diel hatching time may determine the behavioural phenotype of an individual.

Review: Use of Electrophysiological Techniques to Study Visual Functions of Aquatic Organisms.

The light environments of natural water sources have specific characteristics. For the majority of aquatic organisms, vision is crucial for predation, hiding from predators, communicating information, and reproduction. Electroretinography (ERG) is a diagnostic method used for assessing visual function. An electroretinogram records the comprehensive potential response of retinal cells under light stimuli and divides it into several components. Unique wave components are derived from different retinal cells, thus retinal function can be determined by analyzing these components. This review provides an overview of the milestones of ERG technology, describing how ERG is used to study visual sensitivity (e.g., spectral sensitivity, luminous sensitivity, and temporal resolution) of fish, crustaceans, mollusks, and other aquatic organisms (seals, sea lions, sea turtles, horseshoe crabs, and jellyfish). In addition, it describes the correlations between visual sensitivity and habitat, the variation of visual sensitivity as a function of individual growth, and the diel cycle changes of visual sensitivity. Efforts to identify the visual sensitivity of different aquatic organisms are vital to understanding the environmental plasticity of biological evolution and for directing aquaculture, marine fishery, and ecosystem management.

The impact of persistent colonization by Vibrio fischeri on the metabolome of the host squid Euprymna scolopes.

Associations between animals and microbes affect not only the immediate tissues where they occur, but also the entire host. Metabolomics, the study of small biomolecules generated during metabolic processes, provides a window into how mutualistic interactions shape host biochemistry. The Hawaiian bobtail squid, Euprymna scolopes, is amenable to metabolomic studies of symbiosis because the host can be reared with or without its species-specific symbiont, Vibrio fischeri In addition, unlike many invertebrates, the host squid has a closed circulatory system. This feature allows a direct sampling of the refined collection of metabolites circulating through the body, a focused approach that has been highly successful with mammals. Here, we show that rearing E. scolopes without its natural symbiont significantly affected one-quarter of the more than 100 hemolymph metabolites defined by gas chromatography mass spectrometry analysis. Furthermore, as in mammals, which harbor complex consortia of bacterial symbionts, the metabolite signature oscillated on symbiont-driven daily rhythms and was dependent on the sex of the host. Thus, our results provide evidence that the population of even a single symbiont species can influence host hemolymph biochemistry as a function of symbiotic state, host sex and circadian rhythm.

Utilization of various forms of nitrogen and expression regulation of transporters in the harmful alga Heterosigma akashiwo (Raphidophyceae).

Nitrogen (N) is an essential nutrient for phytoplankton growth. There is ample evidence that N enrichment promotes harmful algae blooms (HABs) but molecular mechanisms regulating N-nutrient uptake and metabolism are not so clear, especially for the raphidophyte Heterosigma akashiwo, which forms ichthyotoxic HABs in many coastal waters. In this study, the utilization of three different chemical forms of N (nitrate, ammonium, and urea) by H. akashiwo CCMA 369 was investigated in batch culture conditions. Results showed that H. akashiwo grew well on all three N compounds, and the highest cell yield occurred in the NH4+ culture group. Reverse transcription quantitative PCR analysis revealed that the expression of high-affinity NO3- transporter (NIT), NH4+ transporter (AMT) and high-affinity urea active transporter (DUR3), were significantly up-regulated under N-limitation compared to the N-replete control. The mRNA levels of AMT and DUR3 also displayed a clear diel rhythm, with high levels at midnight. In addition, NH4+ addition (5 µM) did not depress the transcript abundance of any of the three N transporters. Compared with the co-occurring immobile diatom Skeletonema costatum, the high expression of AMT in dark period in H. akashiwo is consistent with its diel vertical migration behavior, which may promote N-nutrient acquisition from deeper layers and give advantages for H. akashiwo to form blooms.

Diel flight activity of wild-caught Anopheles farauti (s.s.) and An. hinesorum malaria mosquitoes from northern Queensland, Australia.

BACKGROUND: Species in the Anopheles farauti complex are major malaria vectors in the Asia Pacific region. Anopheline mosquitoes exhibit circadian and diel rhythms in sugar- and blood-feeding (biting), flight activity, oviposition, and in some species, a short-lived dusk/early night associated swarming behaviour during which mating occurs. A behavioural study of wild-caught mosquitoes from Queensland, Australia was conducted to investigate the differences in diel rhythmic flight activity between two cryptic species in several reproductive states. RESULTS: The 24-hour flight activity of individual adult female mosquitoes under light:dark cycle conditions were monitored with a minute-to-minute time resolution using an infrared beam break method. Mosquitoes were analyzed for reproductive state (insemination and parity) and identified to species [An. farauti (s.s.) Laveran and An. hinesorum Schmidt] by PCR analysis. We compared daily total flight activity, timing of activity onset, the peak in early nocturnal activity, and patterns of activity during the scotophase (night). Species-specific differences between An. farauti and An. hinesorum were observed. Compared to An. farauti, An. hinesorum had an earlier onset of dusk activity, an earlier peak in nocturnal activity, and a higher level of activity at the onset of darkness. Small differences between species were also observed in the pattern of the dusk/early-night bouts of activity. A second nocturnal peak in inseminated nulliparous An. hinesorum was also observed during the middle of the scotophase. CONCLUSIONS: The behavioural differences between these two sympatric species of the An. farauti complex might contribute to subtle differences in habitat adaptation, the timing of host-seeking and/or sugar-feeding activity. This study provides baseline data for analysis of populations of mosquitoes from other geographical regions where these species are malaria vectors, such as in the Solomon Islands and Papua New Guinea. This is important as selective pressures due to long-term use of indoor residual spraying of insecticides and insecticide-treated bed nets are shifting the nocturnal profile of biting behaviour of these vectors to earlier in the night.

Diel Rhythms in Monochamus (Coleoptera: Cerambycidae): Production of and Response to a Male-Produced Aggregation Pheromone.

Recent research has shown that several species in the genus Monochamus, including Monochamus scutellatus scutellatus (Say) and Monochamus notatus (Drury), a close congener, are attracted to an aggregation pheromone, monochamol, but only M. s. scutellatus produces it. Investigations were conducted to determine if there is a diel rhythm in production of monochamol or response to monochamol + host attractants in field trials by M. s. scutellatus and M. notatus to prevent cross attraction with each other. Volatiles were collected from males and females of M. s. scutellatus and M. notatus every 8 h and analyzed by coupled gas chromatography-mass spectrometry. Traps baited with monochamol and the host volatiles α-pinene and ethanol were checked every 8 h in field tests. Only male M. s. scutellatus produced the pheromone, and did so in similar quantities during each 8-h time interval assessed, suggesting there is no diel rhythm in pheromone production. In field tests conducted in early to mid-July, significantly more M. s. scutellatus were captured during morning hours, and significantly more M. notatus were caught during afternoon or early evening hours, suggesting temporal partitioning of flight or in their response to the pheromone lures when both species are present. A temporal switch occurred later in the seasonal flight period (mid-August) when densities of M. s. scutellatus were low; all M. notatus and only one M. s. scutellatus were caught during morning hours. This suggests a temporal separation in mate location behavior when both species are abundant and using the same host to avoid interspecific mating.

Sources of diel variation in energetic physiology in an Arctic-breeding, diving seaduck.

Diel variation in baseline glucocorticoid (GC) secretion influences energetics and foraging behaviors. In temperate breeding, diurnal vertebrates, studies have shown that daily patterns of baseline GC secretion are influenced by environmental photoperiod, with baseline GCs peaking prior to sunrise to stimulate waking and foraging behaviors. Measures of physiological energy acquisition are also expected to peak in response to foraging activity, but their relationship to GC levels have not been well studied. In contrast to temperate breeding species, virtually nothing is known about diel GC and energetic metabolite secretion in Arctic breeding species, which experience almost constant photoperiods in spring and summer. Using a ten-year dataset, we examined the daily, 24-h pattern of baseline corticosterone (CORT) and triglyceride (TRIG) secretion in approximately 800 female pre-breeding Arctic-nesting common eiders (Somateria mollissima). We related these traits to environmental photoperiod and to tidal cycle. In contrast to temperate breeding species, we found that that neither time of day nor tidal trend predicted diel variation in CORT or TRIG secretion in Arctic-breeding eiders. Given the narrow window of opportunity for breeding in polar regions, we suggest that eiders must decouple their daily foraging activity from light and tidal cycles if they are to accrue sufficient energy for successful breeding. As CORT is known to influence foraging behavior, the absence of a distinct diel pattern of CORT secretion may therefore be an adaptation to optimize reproductive investment and likelihood for success in some polar-breeding species.

Time-of-day specific changes in metabolic detoxification and insecticide resistance in the malaria mosquito Anopheles gambiae.

Mosquitoes exhibit ∼24 h rhythms in physiology and behavior, regulated by the cooperative action of an endogenous circadian clock and the environmental light:dark cycle. Here, we characterize diel (observed under light:dark conditions) time-of-day changes in metabolic detoxification and resistance to insecticide challenge in Anopheles gambiae mosquitoes. A better understanding of mosquito chronobiology will yield insights into developing novel control strategies for this important disease vector. We have previously identified >2000 rhythmically expressed An. gambiae genes. These include metabolic detoxification enzymes peaking at various times throughout the day. Especially interesting was the identification of rhythmic genes encoding enzymes capable of pyrethroid and/or DDT metabolism (CYP6M2, CYP6P3, CYP6Z1, and GSTE2). We hypothesized that these temporal changes in gene expression would confer time-of-day specific changes in metabolic detoxification and responses to insecticide challenge. An. gambiae mosquitoes (adult female Pimperena and Mali-NIH strains) were tested by gene expression analysis for diel rhythms in key genes associated with insecticidal resistance. Biochemical assays for total GST, esterase, and oxidase enzymatic activities were undertaken on time-specific mosquito head and body protein lysates. To determine for rhythmic susceptibility to insecticides by survivorship, mosquitoes were exposed to DDT or deltamethrin across the diel cycle. We report the occurrence of temporal changes in GST activity in samples extracted from the body and head with a single peak at late-night to dawn, but no rhythms were detected in oxidase or esterase activity. The Pimperena strain was found to be resistant to insecticidal challenge, and subsequent genomic analysis revealed the presence of the resistance-conferring kdr mutation. We observed diel rhythmicity in key insecticide detoxification genes in the Mali-NIH strain, with peak phases as previously reported in the Pimperena strain. The insecticide sensitive Mali-NIH strain mosquitoes exhibited a diel rhythm in survivorship to DDT exposure and a bimodal variation to deltamethrin challenge. Our results demonstrate rhythms in detoxification and pesticide susceptibility in An. gambiae mosquitoes; this knowledge could be incorporated into mosquito control and experimental design strategies, and contributes to our basic understanding of mosquito biology.