Larval Eye Research Articles

A Case of Larval Oriental Eye Worm, Thelazia callipaeda, Infection Overlooked in Routine Ophthalmic Practice

A Case of Larval Oriental Eye Worm, Thelazia callipaeda, Infection Overlooked in Routine Ophthalmic Practice

Single-cell sequencing suggests a conserved function of Hedgehog-signalling in spider eye development

BackgroundSpiders evolved different types of eyes, a pair of primary eyes that are usually forward pointing, and three pairs of secondary eyes that are typically situated more posterior and lateral on the spider’s head. The best understanding of arthropod eye development comes from the vinegar fly Drosophila melanogaster, the main arthropod model organism, that also evolved different types of eyes, the larval eyes and the ocelli and compound eyes of the imago. The gene regulatory networks that underlie eye development in this species are well investigated revealing a conserved core network, but also show several differences between the different types of eyes. Recent candidate gene approaches identified a number of conserved genes in arthropod eye development, but also revealed crucial differences including the apparent lack of some key factors in some groups of arthropods, including spiders.ResultsHere, we re-analysed our published scRNA sequencing data and found potential key regulators of spider eye development that were previously overlooked. Unlike earlier research on this topic, our new data suggest that Hedgehog (Hh)-signalling is involved in eye development in the spider Parasteatoda tepidariorum. By investigating embryonic gene expression in representatives of all main groups of spiders, we demonstrate that this involvement is conserved in spiders. Additionally, we identified genes that are expressed in the developing eyes of spiders, but that have not been studied in this context before.ConclusionOur data show that single-cell sequencing represents a powerful method to gain deeper insight into gene regulatory networks that underlie the development of lineage-specific organs such as the derived set of eyes in spiders. Overall, we gained deeper insight into spider eye development, as well as the evolution of arthropod visual system formation.

EyaHOST, a modular genetic system for investigation of intercellular and tumor-host interactions in Drosophila melanogaster.

* eyes absent , eye disc-specific enhancer drives clonal KD recombinase flip-out activated QF2 expression in the larval eye epithelium for simultaneous QUAS-driven gain and loss-of-function analysis of gene function. *Clones are visualized by QUAS-tagBFP or QUAS-eGFP facilitating analysis of existing fluorescent reporters.*The GAL4-UAS system and existing genome-wide genetic tools are released to independently manipulate any cell population in the animal for cell biology, intercellular or inter-organ analysis for developmental, physiological, or disease model analysis.*Fly avatars for tumor-host interaction studies with multiple organs allow live monitoring and manipulation of tumors and organs in translucent larva.

A single cell RNA sequence atlas of the early Drosophila larval eye

The Drosophila eye has been an important model to understand principles of differentiation, proliferation, apoptosis and tissue morphogenesis. However, a single cell RNA sequence resource that captures gene expression dynamics from the initiation of differentiation to the specification of different cell types in the larval eye disc is lacking. Here, we report transcriptomic data from 13,000 cells that cover six developmental stages of the larval eye. Our data show cell clusters that correspond to all major cell types present in the eye disc ranging from the initiation of the morphogenetic furrow to the differentiation of each photoreceptor cell type as well as early cone cells. We identify dozens of cell type-specific genes whose function in different aspects of eye development have not been reported. These single cell data will greatly aid research groups studying different aspects of early eye development and will facilitate a deeper understanding of the larval eye as a model system.

Integrative genomic analyses reveal putative cell type-specific targets of the Drosophila ets transcription factor Pointed

The Ets domain transcription factors direct diverse biological processes throughout all metazoans and are implicated in development as well as in tumor initiation, progression and metastasis. The Drosophila Ets transcription factor Pointed (Pnt) is the downstream effector of the Epidermal growth factor receptor (Egfr) pathway and is required for cell cycle progression, specification, and differentiation of most cell types in the larval eye disc. Despite its critical role in development, very few targets of Pnt have been reported previously. Here, we employed an integrated approach by combining genome-wide single cell and bulk data to identify putative cell type-specific Pnt targets. First, we used chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) to determine the genome-wide occupancy of Pnt in late larval eye discs. We identified enriched regions that mapped to an average of 6,941 genes, the vast majority of which are novel putative Pnt targets. Next, we integrated ChIP-seq data with two other larval eye single cell genomics datasets (scRNA-seq and snATAC-seq) to reveal 157 putative cell type-specific Pnt targets that may help mediate unique cell type responses upon Egfr-induced differentiation. Finally, our integrated data also predicts cell type-specific functional enhancers that were not reported previously. Together, our study provides a greatly expanded list of putative cell type-specific Pnt targets in the eye and is a resource for future studies that will allow mechanistic insights into complex developmental processes regulated by Egfr signaling.

Live imaging of photoreceptor outer segment phagocytosis in zebrafish

Retina depends on retinal pigment epithelium (RPE) for its survival. To maintain normal functionality, photoreceptors renew 10% of their outer segments every day and this process requires efficient phagocytosis of the aged photoreceptor fragments by the RPE. Phagocytosis is a dynamic process with strong circadian regulation, but detailed understanding on the regulation of the process and differences between photoreceptor types is presently inadequate. Previous studies have been carried out mainly with in vitro models and cell cultures, and therefore, the dynamics of the phagocytosis process in the context of a living animal has remained elusive. Here, we present an approach that utilizes transgenic zebrafish lines with fluorescently tagged photoreceptor and RPE cells for in vivo imaging of their interactive functions in a living animal.Zebrafish is one of the most accessible animal models to study retinal physiology in vivo and currently the only vertebrate model that can be used to image retinal tissue dynamics in real time. By utilizing already existing low‐pigmented zebrafish lines and microinjecting plasmids coding for photoreceptor and RPE cell membrane targeted tags, we have created low‐pigmented fish with fluorescent RPE cells and the outer segments of the different photoreceptor types. Here, we first show with histological sections how the plasmids are expressed in the correct photoreceptor types and RPE cells in larval zebrafish eyes. Then, we present our in vivo imaging approach where we anesthetise and mount the zebrafish to a confocal microscope. Our results using time‐lapse imaging demonstrate how we can follow the interaction between photoreceptors and RPE in live animals in resolution that enables detecting the movement of photoreceptor outer segment fragments in the subretinal space. Taken together, our study introduces a new approach to investigate the dynamics of phagocytosis of different photoreceptor types by the RPE in real time in a living animal.

Transcriptome-based approach to identify mechanisms underlying locomotor abnormality induced by decabromodiphenyl ethane in zebrafish larvae

The brominated flame retardant decabromodiphenyl ethane (DBDPE) has been extensively used following restrictions on BDE-209 and thus, been frequently detected in aquatic environment. However, information on impact of DBDPE on fish development and the potential mechanisms remains scarce. In present study, developing zebrafish were employed as a study model. Embryos were exposed until 5 d to DBDPE at concentrations of 0, 3, 30, and 300 μg/L, following which the impact on larval development was investigated. DBDPE bioaccumulation and locomotor hyperactivity were observed in developing zebrafish exposed to DBDPE. Transcriptome and bioinformatics analyses indicated that pathways associated with cardiac muscle contraction and retinol metabolism were notably affected. The mechanisms of DBDPE to induce locomotor abnormality were further investigated by analyzing levels of retinol and retinol metabolites, eye and heart histology, heart rates, and ATPase activity. Our results indicate that locomotor hyperactivity observed in larvae exposed to DBDPE results from abnormal heartbeat, which in turn is attributable to inhibition of Na+/K+-ATPase activity. Furthermore, DBDPE did not change larval eye histology and contents of retinoid (retinol, retinal, and retinoic acid). This study provides insight into the mechanisms underlying DBDPE-induced developmental toxicity and highlights the need for addressing the environmental risks for aquatic organisms.

The effect of continuous lighting on the rearing of yellowfin tuna (Thunnus albacares) larvae

Yellowfin tuna (Thunnus albacares) has important economic value in the world market. This study aims to determine the effect of continuous lighting on survival, growth, stomach contents, and histology of the eye of larvae. Larval rearing was conducted on 6 concrete tanks with 60 m3 in volume that support the water circulation and aeration system with an initial stocking of 20 eggs/liter. The treatments tested were A: Larval rearing by a 40-watt fluorescent lamp (TL) which was positioned 1 m above the surface of the water surface for 24 hours and B: Without lights (control). Each treatment used 3 repetitions and data analysis with a t-test. The parameters observed were larval survival and growth stomach content and eyes histology. The results showed that the rearing of larvae in treatment B can with a survival of 0.26±0.0231% and a final total length of 3 6.2 ± 7.3 mm which is significantly higher (P<0.05) compared to treatment A (0.10±0.057% and 24.3 ± 5.5 mm). From the observation of stomach contents at night in treatment B still eats rotifers while in treatment A stomach is empty. The inner nuclear layer cell arrangement of larval eyes is more tenuous than control. From these results, it can be concluded that the provision of continuous lighting treatment can improve the survival, growth, stomach content, and vision ability of larvae.

Exploring the molecular makeup of support cells in insect camera eyes

Animals typically have either compound eyes, or camera-type eyes, both of which have evolved repeatedly in the animal kingdom. Both eye types include two important kinds of cells: photoreceptor cells, which can be excited by light, and non-neuronal support cells (SupCs), which provide essential support to photoreceptors. At the molecular level deeply conserved genes that relate to the differentiation of photoreceptor cells have fueled a discussion on whether or not a shared evolutionary origin might be considered for this cell type. In contrast, only a handful of studies, primarily on the compound eyes of Drosophila melanogaster, have demonstrated molecular similarities in SupCs. D. melanogaster SupCs (Semper cells and primary pigment cells) are specialized eye glia that share several molecular similarities with certain vertebrate eye glia, including Müller glia. This led us to question if there could be conserved molecular signatures of SupCs, even in functionally different eyes such as the image-forming larval camera eyes of the sunburst diving beetle Thermonectus marmoratus. To investigate this possibility, we used an in-depth comparative whole-tissue transcriptomics approach. Specifically, we dissected the larval principal camera eyes into SupC- and retina-containing regions and generated the respective transcriptomes. Our analysis revealed several common features of SupCs including enrichment of genes that are important for glial function (e.g. gap junction proteins such as innexin 3), glycogen production (glycogenin), and energy metabolism (glutamine synthetase 1 and 2). To evaluate similarities, we compared our transcriptomes with those of fly (Semper cells) and vertebrate (Müller glia) eye glia as well as respective retinas. T. marmoratus SupCs were found to have distinct genetic overlap with both fly and vertebrate eye glia. These results suggest that T. marmoratus SupCs are a form of glia, and like photoreceptors, may be deeply conserved.

Larval Body Shape and Eye Size Variation in Japanese Toads (Anura: Bufonidae: Bufo): An Ecological Implication for Lentic and Lotic Habitats

Larval Body Shape and Eye Size Variation in Japanese Toads (Anura: Bufonidae: Bufo): An Ecological Implication for Lentic and Lotic Habitats

A single cell genomics atlas of the Drosophila larval eye reveals distinct photoreceptor developmental timelines

The Drosophila eye is a powerful model system to study the dynamics of cell differentiation, cell state transitions, cell maturation, and pattern formation. However, a high-resolution single cell genomics resource that accurately profiles all major cell types of the larval eye disc and their spatiotemporal relationships is lacking. Here, we report transcriptomic and chromatin accessibility data for all known cell types in the developing eye. Photoreceptors appear as strands of cells that represent their dynamic developmental timelines. As photoreceptor subtypes mature, they appear to assume a common transcriptomic profile that is dominated by genes involved in axon function. We identify cell type maturation genes, enhancers, and potential regulators, as well as genes with distinct R3 or R4 photoreceptor specific expression. Finally, we observe that the chromatin accessibility between cones and photoreceptors is distinct. These single cell genomics atlases will greatly enhance the power of the Drosophila eye as a model system.

Mechanism of thyroid hormone and its structurally similar contaminant bisphenol S exposure on retinoid metabolism in zebrafish larval eyes

The photoreceptor necessitates the retinoids metabolism processes in visual cycle pathway to regenerate visual pigments and sustain vision. Bisphenol S (BPS), with similar structure of thyroid hormone (TH), was reported to impair the light-sensing function of zebrafish larvae via disturbing TH-thyroid hormone receptor β (TRβ) signaling pathway. However, it remains unknown whether TRβ could modulate the toxicity of BPS on retinoid metabolism in visual cycle. This study showed that BPS diminished the optokinetic response of zebrafish larvae and had a stimulative effect on all-trans-retinoic acid (atRA) metabolism, like exogenous T3 exposure. By modulating CYP26A1 and TRβ expression, it was found that CYP26A1 played a crucial role in catalyzing oxidative metabolism of atRA and retinoids regeneration in visual cycle, and TRβ mediated cyp26a1 expression in zebrafish eyes. Similar with 10 nM T3 treatment, cyp26a1 expression could be induced by BPS in the presence of TRβ. Further, in CYP26A1 and TRβ- deficient eyes, 100 μg/L BPS could no longer promote atRA metabolism, or decrease the all-trans-retinol and 11-cis retinal contents in visual cycle, demonstrating that BPS exposure disturbed CYP26A1-mediated visual retinoids metabolism via TRβ. Overall, this study highlights the role of TRβ in mediating the retinoids homeostasis disruption caused by BPS, and provides new clues for exploring molecular targets of visual toxicity under pollutants stress.

Mechanistic insight into the adverse outcome of tire wear and road particle leachate exposure in zebrafish (Danio rerio) larvae

Tire wear particles (TWP) have become the major microplastic pollution in China. Road runoff containing TWP leachate can decrease the eye size and even induced mortality in the aquatic organisms. However, the toxic mechanism of TWP and road particles (RP) leachate on aquatic organisms is still unclear. In this study, the zebrafish embryos were exposed to TWP or RP leachate for 5 days at both environmental relevant and high concentrations. The adverse outcome pathways (AOPs) were screened from individual to molecular levels. The morphological and behavioral analysis demonstrated that the leachate exposure mainly impaired the eye development of zebrafish larvae and inhibited the larval swim behavior and phototactic response, which are the adverse outcomes. The phototransduction modulated by zebrafish retina was significantly down-regulated through transcriptomics and metabolomics analysis. The eye histopathological analysis showed that the decreased thickness of the retinal outer nuclear layer (ONL) and retinal pigmented epithelium (RPE) after leachate exposure were caused by the decreased photoreceptor cells. Moreover, the expression of NR2E3 and TPO genes showed concentration-dependent down-regulation after leachate exposure. The inhibition of photoreceptor cell proliferation was identified as the main reason for photoreceptor cell decrease in zebrafish larval eye. This study, for the first time, uncovered the underlying toxic mechanism of TWP and RP on zebrafish larval eyes.

Interommatidial cells build a tensile collagen network during Drosophila retinal morphogenesis

Drosophila compound eye morphogenesis transforms a simple epithelium into an approximate hollow hemisphere comprised of ∼700 ommatidia, packed as tapering hexagonal prisms between a rigid external array of cuticular lenses and a parallel, rigid internal floor, the fenestrated membrane (FM). Critical to vision, photosensory rhabdomeres are sprung between these two surfaces, grading their length and shape accurately across the eye and aligning them to the optical axis. Using fluorescently tagged collagen and laminin, we show that that the FM assembles sequentially, emerging in the larval eye disc in the wake of the morphogenetic furrow as the original collagen-containing basement membrane (BM) separates from the epithelial floor and is replaced by a new, laminin-rich BM, which advances around axon bundles of newly differentiated photoreceptors as they exit the retina, forming fenestrae in this new, laminin-rich BM. In mid-pupal development, the interommatidial cells (IOCs) autonomously deposit collagen at fenestrae, forming rigid, tension-resisting grommets. In turn, stress fibers assemble in the IOC basal endfeet, where they contact grommets at anchorages mediated by integrin linked kinase (ILK). The hexagonal network of IOC endfeet tiling the retinal floor couples nearest-neighbor grommets into a supracellular tri-axial tension network. Late in pupal development, IOC stress fiber contraction folds pliable BM into a hexagonal grid of collagen-stiffened ridges, concomitantly decreasing the area of convex FM and applying essential morphogenetic longitudinal tension to rapidly growing rhabdomeres. Together, our results reveal an orderly program of sequential assembly and activation of a supramolecular tensile network that governs Drosophila retinal morphogenesis.

Physiological and behavioral evidence for multiple spectral channels in the larval stomatopod visual system.

Larval stomatopods have generally been described as having a typical larval crustacean compound eye, which lacks the visual pigment diversity and morphological specializations of the well-studied stomatopod adult eye. However, recent work has suggested that larval stomatopod eyes are more complex than previously described. In this study we provide physiological and behavioral evidence of at least three distinct photoreceptor classes in three species of larval stomatopods: Gonodactylellus n. sp., Gonodactylaceus falcatus, and Pullosquilla n. sp. First, electroretinogram recordings were used to measure the spectral sensitivity of each species. Evidence for at least three spectral classes were identified in each: a UV peaking at 340-376 nm, a short wavelength blue peaking at 455-464 nm, and a long wavelength orange peaking at 576-602 nm. Next, the behavioral response to light was investigated. We found that each species demonstrated positive phototactic responses to monochromatic stimuli across the ultraviolet-visible spectrum. In wavelength preference trials, distinct preferences among species were identified when different colored light stimuli were presented simultaneously. All species displayed a strong response to the UV stimulus, as well as responses to blue and orange stimuli, although at different response strengths, but no response to green. The results of this study demonstrate that larval stomatopods not only have multiple physiologically active spectral classes, but they also display clear and distinct responses to colors across the spectrum. We propose that the spectral classes demonstrated in each are related to visually guided ecological tasks of the larvae, which may differ between species.

Peripodial adherens junctions regulate Ajuba-Yorkie signaling to preserve fly eye morphology.

The Drosophila eye develops from the larval eye disc, a flattened vesicle comprised of continuous retinal and peripodial epithelia (PE). The PE is an epithelium that plays a supporting role in retinal neurogenesis, but gives rise to cuticle in the adult. We report here that the PE is also necessary to preserve the morphology of the retinal epithelium. Depletion of the adherens junction (AJ) components β-Catenin (β-Cat), DE-Cadherin or α-Catenin from the PE leads to altered disc morphology, characterized by retinal displacement (RDis); so too does loss of the Ajuba protein Jub, an AJ-associated regulator of the transcriptional coactivator Yorkie (Yki). Restoring AJs or overexpressing Yki in β-Cat deficient PE results in suppression of RDis. Additional suppressors of AJ-dependent RDis include knockdown of Rho kinase (Rok) and Dystrophin (Dys). Furthermore, knockdown of βPS integrin (Mys) from the PE results in RDis, while overexpression of Mys can suppress RDis induced by the loss of β-Cat. We thus propose that AJ-Jub-Yki signaling in PE cells regulates PE cell contractile properties and/or attachment to the extracellular matrix to promote normal eye disc morphology.

Investigation of the ultrastructures and retinal arrangements of larval stomatopod eyes

Though the transparent apposition eyes of larval stomatopod crustaceans lack most of the unique retinal specializations known from their adult counterparts, increasing evidence suggests that these tiny pelagic organisms possess their own version of retinal complexity. In this paper, we examined the structural organization of larval eyes in six species of stomatopod crustaceans across three stomatopod superfamilies using transmission electron microscopy. The primary focus was to examine retinular cell arrangement of the larval eyes and characterize the presence of an eighth retinular cell (R8), which is typically responsible for UV vision in crustaceans. For all species investigated, we identified R8 photoreceptor cells positioned distal to the main rhabdom of R1-7 cells. This is the first evidence that R8 photoreceptor cells exist in larval stomatopod retinas, and among the first identified in any larval crustacean. Considering recent studies that identified UV sensitivity in larval stomatopods, we propose that this sensitivity is driven by this putative R8 photoreceptor cell. Additionally, we identified a potentially unique crystalline cone structure in each of the species examined, the function of which is still not understood.

Buffered EGFR signaling regulated by spitz-to-argos expression ratio is a critical factor for patterning the Drosophila eye.

The Epidermal Growth Factor Receptor (EGFR) signaling pathway plays a critical role in regulating tissue patterning. Drosophila EGFR signaling achieves specificity through multiple ligands and feedback loops to finetune signaling outcomes spatiotemporally. The principal Drosophila EGF ligand, cleaved Spitz, and the negative feedback regulator, Argos are diffusible and can act both in a cell autonomous and non-autonomous manner. The expression dose of Spitz and Argos early in photoreceptor cell fate determination has been shown to be critical in patterning the Drosophila eye, but the exact identity of the cells expressing these genes in the larval eye disc has been elusive. Using single molecule RNA Fluorescence in situ Hybridization (smFISH), we reveal an intriguing differential expression of spitz and argos mRNA in the Drosophila third instar eye imaginal disc indicative of directional non-autonomous EGFR signaling. By genetically tuning EGFR signaling, we show that rather than absolute levels of expression, the ratio of expression of spitz-to-argos to be a critical determinant of the final adult eye phenotype. Proximate effects on EGFR signaling in terms of cell cycle and differentiation markers are affected differently in the different perturbations. Proper ommatidial patterning is robust to thresholds around a tightly maintained wildtype spitz-to-argos ratio, and breaks down beyond. This provides a powerful instance of developmental buffering against gene expression fluctuations.

Bisphenol S disrupts opsins gene expression and impairs the light-sensing function via antagonizing TH-TRβ signaling pathway in zebrafish larvae.

Bisphenol S (BPS) is extensively used in "bisphenol A-free" products such as baby bottles. Although the visual toxicity of BPS has been reported, the underlying mechanism was largely unknown. In the present study, zebrafish were exposed to 0, 4 and 400nM BPS from 2h post-fertilization (hpf) to 120 hpf to further explore the thyroid disruption mechanism underlying the BPS induced impairment of visual function. The results showed that BPS decreased T3 levels in larval eyes, induced retinal expression of thyroid hormone receptor β (TRβ), and thereby down-regulated the expression of TH-mediated opsin genes (opn1lw1, opn1lw2, opn1mw1, opn1mw2, opn1mw3, and opn1sw2) and impaired subsequent phototransduction pathways, leading to decreased visually mediated phototactic response and body color adaptation but stimulated visual motor response (VMR). Combining exposure of exogenous T3 or 1-850 (antagonist for TRβ) with BPS could partly compensate the inhibited expression of opsin genes (opn1mw2, opn1lw1, and opn1lw2) and alleviate the hyperactivity of larval VMR caused by BPS alone, suggesting that BPS disrupted the opsins expression and also light-sensing function via antagonizing TH-TRβ signaling pathway. This study underlined the importance of TH signaling in regulating the proper vision and proposed a novel mechanism for the visual toxicity of BPS.

Mayflies see the world in a different light

Everyone gains new perspectives as they grow up, but insects that go through metamorphosis see with a completely different set of eyes as adults. Mayflies spend most of their lives under water as larvae before emerging as adults for one final day of speed dating in a last-ditch effort to produce the next generation. In order to compare how mayflies navigate the two different visual landscapes in which they live, Ádám Egri with Ádám Mészáros and György Kriska at the Eötvös Loránd Research Network Centre for Ecological Research, Hungary, measured how mayflies’ eyes respond to different wavelengths of light and compared what they found to the light levels where the insects tend to congregate.Adult mayflies don't have much time to travel so they spend most of their airborne life mingling around the local neighborhood. By reviewing footage from webcam videos, the researchers determined that Ephoronvirgo mayflies typically swarm in the evening when the sun elevation is low, about –15 to –7 deg below the horizon. They then measured the visual environment when the sun is at different elevations with a spectrophotometer, which measures the strength of light across a range of wavelengths. It turned out that light levels when the adults congregate at dusk are low and the majority of the light is made up of short wavelength colours, closer to ultraviolet. In contrast, the water where the larvae grow up is murky, so longer wavelengths like visible green light are better able to permeate the darkness. These differences suggest that the insects’ eyes may be adapted to distinct environments throughout their life cycle.In order to measure what the insects can see, the team collected larval and adult wild mayflies to test their vision in the lab. Light is sensed when photons are absorbed by specialized cells in the eye that communicate with the brain via electrical signals. The researchers measured these electrical signals with a fine electrode probe while shining lights of different wavelengths onto the insects’ eyes. The adult mayfly's eyes were less sensitive to light that is visible to humans and instead picked up on short ultraviolet wavelengths that are invisible to us. Given that the adults emerge in the evening and die the following day, the reduced sensitivity to visual light suits the insects. However, the larvae living in muddy riverbeds sense light and darkness through simple eyes called ocelli, which absorb photons and send electrical signals to the brain, but can't resolve shapes. The researchers found that larval eyes were also sensitive to UV light, but they were most sensitive to light with longer wavelengths closer to green, which is fitting given the murky world they grow up in.Many insects can see ultraviolet, but these experiments shed light on the green sensitivity of larval ocelli. Although they can't see as well as other animals, the simple eyes of larval mayflies are well matched for the muddy riverbed conditions where they spend most of their lives. As adults, mayflies emerge and face their first and last evening of life above water with a fresh set of eyes that help them find a mate in the dark.