Jan M. Hemmi

In the largest habitat on earth—the deep open ocean—water depth is typically linked to adaptations for camouflage and vision. Recently, it was hypothesized that association behaviour is a primary evolutionary driver of the great eye diversity in a group of crustaceans, the hyperiid amphipods (suborder Hyperiidea). Here, we test that hypothesis by investigating the link between association behaviour (symbiotic or free swimming), depth, eye complexity and camouflage across a broad range of hyperiid diversity. We collated a unique dataset containing observations of 48 of the 80 hyperiid genera from a comprehensive literature review, blackwater scuba photographs and nearly 30 years of submersible observations. By scoring association behaviour in 5759 in situ observations, we show that approximately one third of hyperiid genera have adopted a free-swimming lifestyle. This supports a shift from the current assumption that all hyperiids live in close association with gelatinous animals, to one allowing a free-swimming, predatory lifestyle in many genera. Using phylogenetic generalized linear mixed model (PGLMM) analyses, we show that a free-swimming lifestyle correlates with body transparency and more complex eyes at upper mesopelagic depths. Our results support the hypothesis that a free-swimming lifestyle drives eye complexity while increasing the need for transparency.

The identification of organisms in environmental samples using metabarcoding relies on factors such as taxonomic assignment methods, genetic markers, reference databases and confidence thresholds for taxonomic assignment. Because lineages evolve at different rates, a global threshold (e.g. an unknown sequence is assigned to a species name using a 97% BLAST percent identity threshold) is likely not optimal for accurate taxonomic assignments across all taxa, but no study has systematically evaluated taxon‐specific confidence thresholds. We developed taxon‐specific confidence thresholds for marine eukaryotes to improve the performance of BLAST‐based assignment methods. We tested our approach using published whole community DNA datasets and evaluated factors affecting the accuracy of taxonomic assignments. We provide confidence thresholds for multiple taxonomic levels using the mitochondrial cytochrome c oxidase subunit I (COI) and large ribosomal RNA (lrRNA or 16S rRNA in metazoans) to achieve five false positive error rates (0%, 1%, 5%, 10% and 20%). Additionally, we test two methods to pick the BLAST final hit, the best‐hit and best‐shared (similar to lowest common ancestor) methods. The best‐shared approach to pick the final BLAST hit reduced assignment errors for lower taxonomic levels, and therefore, all subsequent analyses utilized this method. The estimated taxon‐specific thresholds required to achieve reliable assignments at predefined error rates varied among phyla, genetic markers and taxonomic levels. With a 5% error rate, optimized thresholds identified a higher number of sequences at the phylum and family levels than global thresholds. For the genus and species levels, global thresholds identified more sequences to lower taxonomic levels but with an increasing false positive error rate from 14% for family, 21% for genus, to 44% for species. Our results suggest that commonly used global thresholds may be too relaxed and lead to misidentifications for lower taxonomic levels. Using taxon‐optimized thresholds can help define an acceptable error rate (that can be study‐dependent), enable the identification of more unknown sequences at higher taxonomic levels or specific groups, and prevent misclassifications at lower taxonomic levels. We developed an R Shiny application to filter users' BLAST results using predefined error rates and their choice of global or optimized thresholds.

In recent years, virtual reality arenas have become increasingly popular for quantifying visual behaviours. By using the actions of a constrained animal to control the visual scenery, the animal perceives that it is moving through a virtual world. Importantly, as the animal is constrained in space, behavioural quantification is facilitated. Furthermore, using computer‐generated visual scenery allows for identification of visual triggers of behaviour. We created a novel virtual reality arena combining machine vision with the gaming engine Unity. For tethered flight, we enhanced an existing multi‐modal virtual reality arena, MultiMoVR, but tracked wing movements using DeepLabCut‐live (DLC‐live). For tethered walking animals, we used FicTrac to track the motion of a trackball. In both cases, real‐time tracking was interfaced with Unity to control the location and rotation of the tethered animal's avatar in the virtual world. We developed a user‐friendly Unity Editor interface, CAVE, to simplify experimental design and data storage without the need for coding. We show that both the DLC‐live‐Unity and the FicTrac‐Unity configurations close the feedback loop effectively and quickly. We show that closed‐loop feedback reduces behavioural artefacts exhibited by walking crabs in open‐loop scenarios, and that flying Eristalis tenax hoverflies navigate towards virtual flowers in closed loop. We show examples of how the CAVE interface can enable experimental sequencing control including use of avatar proximity to virtual objects of interest. Our results show that combining Unity with machine vision tools provides an easy and flexible virtual reality environment that can be readily adjusted to new experiments and species. This can be implemented programmatically in Unity, or by using our new tool CAVE, which allows users to design new experiments without additional programming. We provide resources for replicating experiments and our interface CAVE via GitHub, together with user manuals and instruction videos, for sharing with the wider scientific community.

The ocean's midwater is a uniquely challenging yet predictable and simple visual environment. The need to see without being seen in this dim, open habitat has led to extraordinary visual adaptations. To understand these adaptations, we compared the morphological and functional differences between the eyes of three hyperiid amphipods— Hyperia galba , Streetsia challengeri and Phronima sedentaria . Combining micro-CT data with computational modelling, we mapped visual field topography and predicted detection distances for visual targets viewed in different directions through mesopelagic depths. Hyperia 's eyes provide a wide visual field optimized for spatial vision over short distances, while Phronima 's and Streetsia 's eyes have the potential to achieve greater sensitivity and longer detection distances using spatial summation. These improvements come at the cost of smaller visual fields, but this loss is compensated for by a second pair of eyes in Phronima and by behaviour in Streetsia . The need to improve sensitivity while minimizing visible eye size to maintain crypsis has likely driven the evolution of hyperiid eye diversity. Our results provide an integrative look at how these elusive animals have adapted to the unique visual challenges of the mesopelagic.

In recent years, virtual reality arenas have become increasingly popular for quantifying visual behaviors. By using the actions of a constrained animal to control the visual scenery, the animal is provided the perception of moving through a simulated environment. As the animal is constrained in space, this allows detailed behavioral quantification. Additionally, as the world is generally computer-generated this allows for mechanistic quantification of visual triggers of behavior. We created a novel virtual arena combining machine vision with the gaming engine Unity. For tethered flight, we enhanced an existing multi-modal virtual reality arena, MultiMoVR (Kaushik et al., 2020) but tracked hoverfly wing movements using DeepLabCut-live (DLC-live, Kane et al., 2020). For trackball experiments, we recorded the motion of a ball that a tethered crab was walking on using FicTrac (Moore et al., 2014). In both cases, real-time tracking was interfaced with Unity to control the movement of the tethered animals’ avatars in the virtual world. We developed a user-friendly Unity Editor interface, CAVE, to simplify experimental design and data storage without the need for coding. We show that both the DLC-live-Unity and the FicTrac-Unity configurations close the feedback loop effectively with small delays, less than 50 ms. Our FicTrac-Unity integration highlighted the importance of closed-loop feedback by reducing behavioral artifacts exhibited by the crabs in open-loop scenarios. We show that Eristalis tenax hoverflies, using the DLC-live-Unity integration, navigate towards flowers. The effectiveness of our CAVE interface is shown by implementing experimental sequencing control based on avatar proximity to virtual structures. Our results show that combining Unity with machine vision tools such as DLC-live and FicTrac provides an easy and flexible virtual reality (VR) environment that can be readily adjusted to new experiments and species. This can be implemented programmatically in Unity, or by using our new tool CAVE, which allows users to design and implement new experiments without programming in code. We provide resources for replicating experiments and our interface CAVE via GitHub, together with user manuals and instruction videos, for sharing with the wider scientific community.

Most animals rely on visual information for a variety of everyday tasks. The information available to a visual system depends in part on its spatial resolving power and contrast sensitivity. Because of their competing demands for physical space within an eye, these traits cannot simultaneously be improved without increasing overall eye size. The contrast sensitivity function is an integrated measure of visual performance that measures both resolution and contrast sensitivity. Its measurement helps us identify how different species have made a trade-off between contrast sensitivity and spatial resolution. It further allows us to identify the evolutionary drivers of sensory processing and visually mediated behaviour. Here, we measured the contrast sensitivity function of the fiddler crab Gelasimus dampieri using its optokinetic responses to wide-field moving sinusoidal intensity gratings of different orientations, spatial frequencies, contrasts and speeds. We further tested whether the behavioural state of the crabs (i.e. whether crabs are actively walking or not) affects their optokinetic gain and contrast sensitivity. Our results from a group of five crabs suggest a minimum perceived contrast of 6% and a horizontal and vertical visual acuity of 0.4 cyc deg−1 and 0.28 cyc deg−1, respectively, in the crabs' region of maximum optomotor sensitivity. Optokinetic gain increased in moving crabs compared with restrained crabs, adding another example of the importance of naturalistic approaches when studying the performance of animals.

Disruptive markings are common in animal patterns and can provide camouflage benefits by concealing the body's true edges and/or by breaking the surface of the body into multiple depth planes. Disruptive patterns that are accentuated by high contrast borders are most likely to provide false depth cues to enhance camouflage, but studies to date have used visual detection models or humans as predators. We presented three-dimensional-printed moth-like targets to wild bird predators to determine whether: (1) three-dimensional prey with disrupted body surfaces have higher survival than three-dimensional prey with continuous surfaces, (2) two-dimensional prey with disruptive patterns or enhanced edge markings have higher survival than non-patterned two-dimensional prey. We found a survival benefit for three-dimensional prey with disrupted surfaces, and a significant effect of mean wing luminance. There was no evidence that false depth cues provided the same protective benefits as physical surface disruption in three-dimensional prey, perhaps because our treatments did not mimic the complexity of patterns found in natural animal markings. Our findings indicate that disruption of surface continuity is an important strategy for concealing a three-dimensional body shape.

Adolescence is a critical period of postnatal development characterized by social, emotional, and cognitive changes. These changes are increasingly understood to depend on white matter development. White matter is highly vulnerable to the effects of injury, including secondary degeneration in regions adjacent to the primary injury site which alters the myelin ultrastructure. However, the impact of such alterations on adolescent white matter maturation is yet to be investigated. To address this, female piebald-virol-glaxo rats underwent partial transection of the optic nerve during early adolescence (postnatal day (PND) 56) with tissue collection two weeks (PND 70) or three months later (PND 140). Axons and myelin in the transmission electron micrographs of tissue adjacent to the injury were classified and measured based on the appearance of the myelin laminae. Injury in adolescence impaired the myelin structure in adulthood, resulting in a lower percentage of axons with compact myelin and a higher percentage of axons with severe myelin decompaction. Myelin thickness did not increase as expected into adulthood after injury and the relationship between the axon diameter and myelin thickness in adulthood was altered. Notably, dysmyelination was not observed 2 weeks postinjury. In conclusion, injury in adolescence altered the developmental trajectory, resulting in impaired myelin maturation when assessed at the ultrastructural level in adulthood.

Vision in the midwater of the open ocean requires animals to perform visual tasks quite unlike those of any other environment. These tasks consist of detecting small, low contrast objects and point sources against a relatively dim and uniform background. Deep-sea animals have evolved many extraordinary visual adaptations to perform these tasks. Linking eye anatomy to specific selective pressures, however, is challenging, not least because of the many difficulties of studying deep-sea animals. Computational modelling of vision, based on detailed morphological reconstructions of animal eyes, along with underwater optics, offers a chance to understand the specific visual capabilities of individual visual systems. Prior to the work presented here, comprehensive models for apposition compound eyes in the mesopelagic, the dominant eye form of crustaceans, were lacking. We adapted a model developed for single-lens eyes and used it to examine how different parameters affect the model’s ability to detect point sources and extended objects. This new model also allowed us to examine spatial summation as a means to improve visual performance. Our results identify a trade-off between increased depth range over which eyes function effectively and increased distance at which extended objects can be detected. This trade-off is driven by the size of the ommatidial acceptance angle. We also show that if neighbouring ommatidia have overlapping receptive fields, spatial summation helps with all detection tasks, including the detection of bioluminescent point sources. By applying our model to the apposition compound eyes ofPhronima, a mesopelagic hyperiid amphipod, we show that the specialisations of the large medial eyes ofPhronimaimprove both the detection of point sources and of extended objects. The medial eyes outperformed the lateral eyes at every modelled detection task. We suggest that the small visual field size ofPhronima’s medial eyes and the strong asymmetry between the medial and lateral eyes reflectPhronima’s need for effective vision across a large depth range and its habit of living inside a barrel. The barrel’s narrow aperture limits the usefulness of a large visual field and has allowed a strong asymmetry between the medial and lateral eyes. The model provides a useful tool for future investigations into the visual abilities of apposition compound eyes in the deep sea.

Selective attention, the ability to focus on a specific stimulus and suppress distractions, plays a fundamental role for animals in many contexts, such as mating, feeding, and predation. Within natural environments, animals are often confronted with multiple stimuli of potential importance. Such a situation significantly complicates the decision-making process and imposes conflicting information on neural systems. In the context of predation, selectively attending to one of multiple threats is one possible solution. However, how animals make such escape decisions is rarely studied. A previous field study on the fiddler crab,Gelasimus dampieri, provided evidence of selective attention in the context of escape decisions. To identify the underlying mechanisms that guide their escape decisions, we measured the crabs’ behavioural and neural responses to either a single, or two simultaneously approaching looming stimuli. The two stimuli were either identical or differed in contrast to represent different levels of threat certainty. Although our behavioural data provides some evidence that crabs perceive signals from both stimuli, we show that both the crabs and their looming-sensitive neurons almost exclusively respond to only one of two simultaneous threats. The crabs’ body orientation played an important role in their decision about which stimulus to run away from. When faced with two stimuli of differing contrasts, both neurons and crabs were much more likely to respond to the stimulus with the higher contrast. Our data provides evidence that the crabs’ looming-sensitive neurons play an important part in the mechanism that drives their selective attention in the context of predation. Our results support previous suggestions that the crabs’ escape direction is calculated downstream of their looming-sensitive neurons by means of a population vector of the looming sensitive neuronal ensemble.