Three-dimensional dental microwear in type-Maastrichtian mosasaur teeth (Reptilia, Squamata).

Mosasaurs (Squamata, Mosasauridae) were large aquatic reptiles from the Late Cretaceous that filled a range of ecological niches within marine ecosystems. The type-Maastrichtian strata (68-66 Ma) of the Netherlands and Belgium preserve remains of five species that seemed to have performed different ecological roles (carnivores, piscivores, durophages). However, many interpretations of mosasaur diet and niche partitioning are based on qualitative types of evidence that are difficult to test explicitly. Here, we apply three-dimensional dental microwear texture analysis (DMTA) to provide quantitative dietary constraints for type-Maastrichtian mosasaurs, and to assess levels of niche partitioning between taxa. DMTA indicates that these mosasaurs did not exhibit neatly defined diets or strict dietary partitioning. Instead, we identify three broad groups: (i) mosasaurs Carinodens belgicus and Plioplatecarpus marshi plotting in the space of modern reptiles that are predominantly piscivorous and/or consume harder invertebrate prey, (ii) Prognathodon saturator and Prognathodon sectorius overlapping with extant reptiles that consume larger amounts of softer invertebrate prey items, and (iii) Mosasaurus hoffmanni spanning a larger plot area in terms of dietary constraints. The clear divide between the aforementioned first two groups in texture-dietary space indicates that, despite our small sample sizes, this method shows the potential of DMTA to test hypotheses and provide quantitative constraints on mosasaur diets and ecological roles.

Dental microwear texture analysis along reptile tooth rows: complex variation with non-dietary variables.

Dental microwear texture analysis (DMTA) is a powerful technique for reconstructing the diets of extant and extinct taxa. Few studies have investigated intraspecific microwear differences along with tooth rows and the influence of endogenous non-dietary variables on texture characteristics. Sampling teeth that are minimally affected by non-dietary variables is vital for robust dietary reconstructions, especially for taxa with non-occlusal (non-chewing) dentitions as no standardized sampling strategies currently exist. Here, we apply DMTA to 13 species of extant reptile (crocodilians and monitor lizards) to investigate intraspecific microwear differences along with tooth rows and to explore the influence of three non-dietary variables on exhibited differences: (i) tooth position, (ii) mechanical advantage, and (iii) tooth aspect ratio. Five species exhibited intraspecific microwear differences. In several crocodilians, the distally positioned teeth exhibited the 'roughest' textures, and texture characteristics correlated with all non-dietary variables. By contrast, the mesial teeth of the roughneck monitor (Varanus rudicollis) exhibited the 'roughest' textures, and texture characteristics did not correlate with aspect ratio. These results are somewhat consistent with how reptiles preferentially use their teeth during feeding. We argue that DMTA has the potential to track mechanical and behavioural differences in tooth use which should be taken into consideration in future dietary reconstructions.

Dental microwear texture analysis as a tool for dietary discrimination in elasmobranchs.

As abundant and widespread predators, elasmobranchs play influential roles in food-web dynamics of marine communities. Clearly, these trophic interactions have significant implications for fisheries management and marine conservation, yet elasmobranch diet is relatively understudied; for the majority of species little or no quantitative dietary data exist. This reflects the difficulties of direct observation of feeding and stomach contents analysis in wild elasmobranchs. Here, by quantifying the 3D surface textures that develop on tooth surfaces as a consequence of feeding, we show that tooth microwear varies with diet in elasmobranchs, providing a new tool for dietary analysis. The technique can be applied to small samples and individuals with no gut contents, and thus offers a way to reduce the impact on wild elasmobranch populations of analysing their dietary ecology, especially relevant in conservation of endangered species. Furthermore, because microwear accumulates over longer periods of time, analysis of texture overcomes the 'snapshot bias' of stomach contents analysis. Microwear texture analysis has the potential to be a powerful tool, complementing existing techniques such as stable isotope analysis, for dietary analysis in living and extinct elasmobranchs.

Dietary diversity and evolution of the earliest flying vertebrates revealed by dental microwear texture analysis.

Pterosaurs, the first vertebrates to evolve active flight, lived between 210 and 66 million years ago. They were important components of Mesozoic ecosystems, and reconstructing pterosaur diets is vital for understanding their origins, their roles within Mesozoic food webs and the impact of other flying vertebrates (i.e. birds) on their evolution. However, pterosaur dietary hypotheses are poorly constrained as most rely on morphological-functional analogies. Here we constrain the diets of 17 pterosaur genera by applying dental microwear texture analysis to the three-dimensional sub-micrometre scale tooth textures that formed during food consumption. We reveal broad patterns of dietary diversity (e.g. Dimorphodon as a vertebrate consumer; Austriadactylus as a consumer of 'hard' invertebrates) and direct evidence of sympatric niche partitioning (Rhamphorhynchus as a piscivore; Pterodactylus as a generalist invertebrate consumer). We propose that the ancestral pterosaur diet was dominated by invertebrates and later pterosaurs evolved into piscivores and carnivores, shifts that might reflect ecological displacements due to pterosaur-bird competition.

Dietary differences in archosaur and lepidosaur reptiles revealed by dental microwear textural analysis.

Reptiles are key components of modern ecosystems, yet for many species detailed characterisations of their diets are lacking. Data currently used in dietary reconstructions are limited either to the last few meals or to proxy records of average diet over temporal scales of months to years, providing only coarse indications of trophic level(s). Proxies that record information over weeks to months would allow more accurate reconstructions of reptile diets and better predictions of how ecosystems might respond to global change drivers. Here, we apply dental microwear textural analysis (DMTA) to dietary guilds encompassing both archosaurian and lepidosaurian reptiles, demonstrating its value as a tool for characterising diets over temporal scales of weeks to months. DMTA, involving analysis of the three-dimensional, sub-micrometre scale textures created on tooth surfaces by interactions with food, reveals that the teeth of reptiles with diets dominated by invertebrates, particularly invertebrates with hard exoskeletons (e.g. beetles and snails), exhibit rougher microwear textures than reptiles with vertebrate-dominated diets. Teeth of fish-feeding reptiles exhibit the smoothest textures of all guilds. These results demonstrate the efficacy of DMTA as a dietary proxy in taxa from across the phylogenetic range of extant reptiles. This method is applicable to extant taxa (living or museum specimens) and extinct reptiles, providing new insights into past, present and future ecosystems.

The phylogenetic signal in tooth wear: What does it mean?

A new study by Fraser et al (2018) urges the use of phylogenetic comparative methods, whenever possible, in analyses of mammalian tooth wear. We are concerned about this for two reasons. First, this recommendation may mislead the research community into thinking that phylogenetic signal is an artifact of some sort rather than a fundamental outcome of the evolutionary process. Secondly, this recommendation may set a precedent for editors and reviewers to enforce phylogenetic adjustment where it may unnecessarily weaken or even directionally alter the results, shifting the emphasis of analysis from common patterns manifested by large clades to rare cases.

Pterosaur dietary hypotheses: a review of ideas and approaches.

Pterosaurs are an extinct group of Mesozoic flying reptiles, whose fossil record extends from approximately 210 to 66 million years ago. They were integral components of continental and marginal marine ecosystems, yet their diets remain poorly constrained. Numerous dietary hypotheses have been proposed for different pterosaur groups, including insectivory, piscivory, carnivory, durophagy, herbivory/frugivory, filter-feeding and generalism. These hypotheses, and subsequent interpretations of pterosaur diet, are supported by qualitative (content fossils, associations, ichnology, comparative anatomy) and/or quantitative (functional morphology, stable isotope analysis) evidence. Pterosaur dietary interpretations are scattered throughout the literature with little attention paid to the supporting evidence. Reaching a robustly supported consensus on pterosaur diets is important for understanding their dietary evolution, and their roles in Mesozoic ecosystems. A comprehensive examination of the pterosaur literature identified 314 dietary interpretations (dietary statement plus supporting evidence) from 126 published studies. Multiple alternative diets have been hypothesised for most principal taxonomic pterosaur groups. Some groups exhibit a high degree of consensus, supported by multiple lines of evidence, while others exhibit less consensus. Qualitative evidence supports 87.3% of dietary interpretations, with comparative anatomy most common (62.1% of total). More speciose groups of pterosaur tend to have a greater range of hypothesised diets. Consideration of dietary interpretations within alternative phylogenetic contexts reveals high levels of consensus between equivalent monofenestratan groups, and lower levels of consensus between equivalent non-monofenestratan groups. Evaluating the possible non-biological controls on apparent patterns of dietary diversity reveals that numbers of dietary interpretations through time exhibit no correlation with patterns of publication (number of peer-reviewed publications through time). 73.8% of dietary interpretations were published in the 21st century. Overall, consensus interpretations of pterosaur diets are better accounted for by non-biological signals, such as the impact of the respective quality of the fossil record of different pterosaur groups on research levels. That many interpretations are based on qualitative, often untestable lines of evidence adds significant noise to the data. More experiment-led pterosaur dietary research, with greater consideration of pterosaurs as organisms with independent evolutionary histories, will lead to more robust conclusions drawn from repeatable results. This will allow greater understanding of pterosaur dietary diversity, disparity and evolution and facilitate reconstructions of Mesozoic ecosystems.

Pigmented anatomy in Carboniferous cyclostomes and the evolution of the vertebrate eye.

The success of vertebrates is linked to the evolution of a camera-style eye and sophisticated visual system. In the absence of useful data from fossils, scenarios for evolutionary assembly of the vertebrate eye have been based necessarily on evidence from development, molecular genetics and comparative anatomy in living vertebrates. Unfortunately, steps in the transition from a light-sensitive 'eye spot' in invertebrate chordates to an image-forming camera-style eye in jawed vertebrates are constrained only by hagfish and lampreys (cyclostomes), which are interpreted to reflect either an intermediate or degenerate condition. Here, we report-based on evidence of size, shape, preservation mode and localized occurrence-the presence of melanosomes (pigment-bearing organelles) in fossil cyclostome eyes. Time of flight secondary ion mass spectrometry analyses reveal secondary ions with a relative intensity characteristic of melanin as revealed through principal components analyses. Our data support the hypotheses that extant hagfish eyes are degenerate, not rudimentary, that cyclostomes are monophyletic, and that the ancestral vertebrate had a functional visual system. We also demonstrate integument pigmentation in fossil lampreys, opening up the exciting possibility of investigating colour patterning in Palaeozoic vertebrates. The examples we report add to the record of melanosome preservation in Carboniferous fossils and attest to surprising durability of melanosomes and biomolecular melanin.

The eyes of Tullimonstrum reveal a vertebrate affinity.

Tullimonstrum gregarium is an iconic soft-bodied fossil from the Carboniferous Mazon Creek Lagerstätte (Illinois, USA). Despite a large number of specimens and distinct anatomy, various analyses over the past five decades have failed to determine the phylogenetic affinities of the 'Tully monster', and although it has been allied to such disparate phyla as the Mollusca, Annelida or Chordata, it remains enigmatic. The nature and phylogenetic affinities of Tullimonstrum have defied confident systematic placement because none of its preserved anatomy provides unequivocal evidence of homology, without which comparative analysis fails. Here we show that the eyes of Tullimonstrum possess ultrastructural details indicating homology with vertebrate eyes. Anatomical analysis using scanning electron microscopy reveals that the eyes of Tullimonstrum preserve a retina defined by a thick sheet comprising distinct layers of spheroidal and cylindrical melanosomes. Time-of-flight secondary ion mass spectrometry and multivariate statistics provide further evidence that these microbodies are melanosomes. A range of animals have melanin in their eyes, but the possession of melanosomes of two distinct morphologies arranged in layers, forming retinal pigment epithelium, is a synapomorphy of vertebrates. Our analysis indicates that in addition to evidence of colour patterning, ecology and thermoregulation, fossil melanosomes can also carry a phylogenetic signal. Identification in Tullimonstrum of spheroidal and cylindrical melanosomes forming the remains of retinal pigment epithelium indicates that it is a vertebrate; considering its body parts in this new light suggests it was an anatomically unusual member of total group Vertebrata.

The impact of taphonomic data on phylogenetic resolution: Helenodora inopinata (Carboniferous, Mazon Creek Lagerstätte) and the onychophoran stem lineage.

BACKGROUND: The origin of the body plan of modern velvet worms (Onychophora) lies in the extinct lobopodians of the Palaeozoic. Helenodora inopinata, from the Mazon Creek Lagerstätte of Illinois (Francis Creek Shale, Carbondale Formation, Middle Pennsylvanian), has been proposed as an intermediate between the "weird wonders" of the Cambrian seas and modern terrestrial predatory onychophorans. The type material of H. inopinata, however, leaves much of the crucial anatomy unknown. RESULTS: Here we present a redescription of this taxon based on more complete material, including new details of the head and posterior portion of the trunk, informed by the results of experimental decay of extant onychophorans. H. inopinata is indeed best resolved as a stem-onychophoran, but lacks several key features of modern velvet worms including, crucially, those that would suggest a terrestrial mode of life. CONCLUSIONS: The presence of H. inopinata in the Carboniferous demonstrates the survival of a Cambrian marine morphotype, and a likely post-Carboniferous origin of crown-Onychophora. Our analysis also demonstrates that taphonomically informed tests of character interpretations have the potential to improve phylogenetic resolution.

Accuracy and precision of silicon based impression media for quantitative areal texture analysis.

Areal surface texture analysis is becoming widespread across a diverse range of applications, from engineering to ecology. In many studies silicon based impression media are used to replicate surfaces, and the fidelity of replication defines the quality of data collected. However, while different investigators have used different impression media, the fidelity of surface replication has not been subjected to quantitative analysis based on areal texture data. Here we present the results of an analysis of the accuracy and precision with which different silicon based impression media of varying composition and viscosity replicate rough and smooth surfaces. Both accuracy and precision vary greatly between different media. High viscosity media tested show very low accuracy and precision, and most other compounds showed either the same pattern, or low accuracy and high precision, or low precision and high accuracy. Of the media tested, mid viscosity President Jet Regular Body and low viscosity President Jet Light Body (Coltène Whaledent) are the only compounds to show high levels of accuracy and precision on both surface types. Our results show that data acquired from different impression media are not comparable, supporting calls for greater standardisation of methods in areal texture analysis.

Decay of velvet worms (Onychophora), and bias in the fossil record of lobopodians.

BACKGROUND: Fossil lobopodians, including animals proposed to have close affinity to modern onychophorans, are crucial to understanding the evolution of the panarthropod body plan and the phylum-level relationships between the ecdysozoan groups. Unfortunately, the key features of their anatomy are un-mineralized and subject to biases introduced during death, decay and preservation, yet the extent to which these fossils have been affected by the processes of post-mortem decay is entirely untested. Recent experimental work on chordates has highlighted a profound bias caused by decay, resulting in the erroneous interpretation of badly decayed specimens as primitive members of a clade (stemward slippage). The degree to which this bias affects organisms other than chordates is unknown. RESULTS: Here we use experimental decay of velvet worms (Onychophora) to examine the importance of decay bias in fossil lobopodians. Although we find stemward slippage is not significant in the interpretation of non-mineralized lobopodian fossils, the affect of decay is far from unbiased. Quantitative analysis reveals significant changes in body proportions during decay, a spectrum of decay resistance across anatomical features, and correlated decay of topologically associated characters. CONCLUSIONS: These results have significant implications for the interpretation of fossil lobopodian remains, demonstrating that features such as body outline and relative proportions are unreliable for taxonomy or phylogenetic reconstruction, unless decay is taken into account. Similarly, the non-independent loss of characters, due to juxtaposition in the body, during decay has the potential to bias phylogenetic analyses of non-biomineralized fossils. Our results are difficult to reconcile with interpretations of highly decay-prone tissues and structures, such as neural tissue, and complex musculature, in recently described Cambrian lobopodians. More broadly, we hypothesize that stemward slippage is unlikely to be a significant factor among the taphonomic biases that have affected organisms where decay-resistant features of the anatomy are rich in phylogenetically informative characters. Conversely, organisms which possess decay-resistant body parts but have informative characters concentrated in decay-prone tissues will be just as liable to bias as those that lack decay-resistant body parts. Further experimental analysis of decay is required to test these hypotheses.

Dietary specializations and diversity in feeding ecology of the earliest stem mammals.

The origin and radiation of mammals are key events in the history of life, with fossils placing the origin at 220 million years ago, in the Late Triassic period. The earliest mammals, representing the first 50 million years of their evolution and including the most basal taxa, are widely considered to be generalized insectivores. This implies that the first phase of the mammalian radiation--associated with the appearance in the fossil record of important innovations such as heterodont dentition, diphyodonty and the dentary-squamosal jaw joint--was decoupled from ecomorphological diversification. Finds of exceptionally complete specimens of later Mesozoic mammals have revealed greater ecomorphological diversity than previously suspected, including adaptations for swimming, burrowing, digging and even gliding, but such well-preserved fossils of earlier mammals do not exist, and robust analysis of their ecomorphological diversity has previously been lacking. Here we present the results of an integrated analysis, using synchrotron X-ray tomography and analyses of biomechanics, finite element models and tooth microwear textures. We find significant differences in function and dietary ecology between two of the earliest mammaliaform taxa, Morganucodon and Kuehneotherium--taxa that are central to the debate on mammalian evolution. Morganucodon possessed comparatively more forceful and robust jaws and consumed 'harder' prey, comparable to extant small-bodied mammals that eat considerable amounts of coleopterans. Kuehneotherium ingested a diet comparable to extant mixed feeders and specialists on 'soft' prey such as lepidopterans. Our results reveal previously hidden trophic specialization at the base of the mammalian radiation; hence even the earliest mammaliaforms were beginning to diversify--morphologically, functionally and ecologically. In contrast to the prevailing view, this pattern suggests that lineage splitting during the earliest stages of mammalian evolution was associated with ecomorphological specialization and niche partitioning.

Morphology of Cambrian lobopodian eyes from the Chengjiang Lagerstätte and their evolutionary significance.

Visual organs are widely distributed throughout the animal kingdom and exhibit a great diversity of morphologies. Compound eyes consisting of numerous visual units (ommatidia) are the oldest preserved visual systems of arthropods, but their origins are obscure and hypothetical models for their evolution have been difficult to test in the absence of unequivocal fossil evidence. Here we reveal the detailed eye structures of well-preserved Early Cambrian lobopodians Luolishania longicruris and Hallucigenia fortis from the Chengjiang Lagerstätte, China. These animals possess a pair of eyes composed of at least two visual units, interpreted as pigment cups. Contrary to previous suggestions that Cambrian lobopodians possessed ocellus-like eyes comparable to those of extant onychophorans, this multi-component structure is more similar to the lateral eyes of arthropods. Morphological comparison and phylogenetic analyses indicate that these lobopodian eyes may represent an early stage in the evolution of the ancestral visual system of euarthropods.

Decay of vertebrate characters in hagfish and lamprey (Cyclostomata) and the implications for the vertebrate fossil record.

The timing and sequence of events underlying the origin and early evolution of vertebrates remains poorly understood. The palaeontological evidence should shed light on these issues, but difficulties in interpretation of the non-biomineralized fossil record make this problematic. Here we present an experimental analysis of decay of vertebrate characters based on the extant jawless vertebrates (Lampetra and Myxine). This provides a framework for the interpretation of the anatomy of soft-bodied fossil vertebrates and putative cyclostomes, and a context for reading the fossil record of non-biomineralized vertebrate characters. Decay results in transformation and non-random loss of characters. In both lamprey and hagfish, different types of cartilage decay at different rates, resulting in taphonomic bias towards loss of 'soft' cartilages containing vertebrate-specific Col2α1 extracellular matrix proteins; phylogenetically informative soft-tissue characters decay before more plesiomorphic characters. As such, synapomorphic decay bias, previously recognized in early chordates, is more pervasive, and needs to be taken into account when interpreting the anatomy of any non-biomineralized fossil vertebrate, such as Haikouichthys, Mayomyzon and Hardistiella.

Non-random decay of chordate characters causes bias in fossil interpretation.

Exceptional preservation of soft-bodied Cambrian chordates provides our only direct information on the origin of vertebrates. Fossil chordates from this interval offer crucial insights into how the distinctive body plan of vertebrates evolved, but reading this pre-biomineralization fossil record is fraught with difficulties, leading to controversial and contradictory interpretations. The cause of these difficulties is taphonomic: we lack data on when and how important characters change as they decompose, resulting in a lack of constraint on anatomical interpretation and a failure to distinguish phylogenetic absence of characters from loss through decay. Here we show, from experimental decay of amphioxus and ammocoetes, that loss of chordate characters during decay is non-random: the more phylogenetically informative are the most labile, whereas plesiomorphic characters are decay resistant. The taphonomic loss of synapomorphies and relatively higher preservation potential of chordate plesiomorphies will thus result in bias towards wrongly placing fossils on the chordate stem. Application of these data to Cathaymyrus (Cambrian period of China) and Metaspriggina (Cambrian period of Canada) highlights the difficulties: these fossils cannot be placed reliably in the chordate or vertebrate stem because they could represent the decayed remains of any non-biomineralized, total-group chordate. Preliminary data suggest that this decay filter also affects other groups of organisms and that 'stem-ward slippage' may be a widespread but currently unrecognized bias in our understanding of the early evolution of a number of phyla.

Quantitative analysis of dental microwear in hadrosaurid dinosaurs, and the implications for hypotheses of jaw mechanics and feeding.

Understanding the feeding mechanisms and diet of nonavian dinosaurs is fundamental to understanding the paleobiology of these taxa and their role in Mesozoic terrestrial ecosystems. Various methods, including biomechanical analysis and 3D computer modeling, have been used to generate detailed functional hypotheses, but in the absence of either direct observations of dinosaur feeding behavior, or close living functional analogues, testing these hypotheses is problematic. Microscopic scratches that form on teeth in vivo during feeding are known to record the relative motion of the tooth rows to each other during feeding and to capture evidence of tooth-food interactions. Analysis of this dental microwear provides a powerful tool for testing hypotheses of jaw mechanics, diet, and trophic niche; yet, quantitative analysis of microwear in dinosaurs has not been attempted. Here, we show that analysis of tooth microwear orientation provides direct evidence for the relative motions of jaws during feeding in hadrosaurid ornithopods, the dominant terrestrial herbivores of the Late Cretaceous. Statistical testing demonstrates that Edmontosaurus teeth preserve 4 distinct sets of scratches in different orientations. In terms of jaw mechanics, these data indicate an isognathic, near-vertical posterodorsal power stroke during feeding; near-vertical jaw opening; and propalinal movements in near anterior and near posterior directions. Our analysis supports the presence of a pleurokinetic hinge, and the straightness and parallelism of scratches indicate a tightly controlled occlusion. The dominance of scratched microwear fabrics suggests that Edmontosaurus was a grazer rather than a browser.

Distinguishing heat from light in debate over controversial fossils.

Fossil organisms offer our only direct insight into how the distinctive body plans of extant organisms were assembled. However, realizing the potential evolutionary significance of fossils can be hampered by controversy over their interpretation. Here, as a guide to evaluating palaeontological debates, we outline the process and pitfalls of fossil interpretation. The physical remains of controversial fossils should be reconstructed before interpreting homologies, and choice of interpretative model should be explicit and justified. Extinct taxa lack characters diagnostic of extant clades because the characters had not yet evolved, because of secondary loss, or because they have rotted away. The latter, if not taken into account, will lead to the spurious assignment of fossils to basally branching clades. Conflicting interpretations of fossils can often be resolved by considering all the steps in the process of anatomical analysis and phylogenetic placement, although we must accept that some fossil organisms are simply too incompletely preserved for their evolutionary significance to be realized.

Correlated evolution and dietary change in fossil stickleback.

The importance of trophic ecology in adaptation and evolution is well known, yet direct evidence that feeding controls microevolution over extended evolutionary time scales, available only from the fossil record, is conspicuously lacking. Through quantitative analysis of tooth microwear, we show that rapid evolutionary change in Miocene stickleback was associated with shifts in feeding, providing direct evidence from the fossil record for changes in trophic niche and resource exploitation driving directional, microevolutionary change over thousands of years. These results demonstrate the potential for tooth microwear analysis to provide powerful insights into trophic ecology during aquatic adaptive radiations.

Quantitative analysis of dental microwear in threespine stickleback: a new approach to analysis of trophic ecology in aquatic vertebrates.

1. The threespine stickleback Gasterosteus aculeatus is an important model organism in studies of genomic and phenotypic evolution, adaptation and speciation. Fossil Gasterosteus offer the potential to test models derived from studies of extant fishes over true evolutionary time-scales. Competition for food resources, for example, plays an important part in stickleback speciation, causing divergence in food gathering traits and ecological character displacement, but it is not possible to test this model in fossils because evidence of diet is almost never preserved. 2. We demonstrate here that quantitative analysis of dental microwear, a technique previously applied only to mammals, provides a reliable guide to the dietary preferences of stickleback. Teeth from stickleback raised under laboratory conditions exhibit microwear patterns that vary systematically according to substrate coarseness and whether fishes feed on Daphnia within the water column, or on chironomid larvae from the bottom. Furthermore, microwear data exhibit a progressive shift in their distribution that tracks differences in experimental feeding treatments. 3. Microwear in wild populations also exhibits a relationship with feeding. In blind assessments of trophic niche based on microwear patterns we were able to correctly assign all but one equivocal population to trophic group. Microwear data from wild stickleback exhibit a shift in distribution comparable with that observed across the range of treatments in the laboratory and these allow populations to be ranked according to the degree to which they approach fully benthic or fully limnetic feeding. 4. Our results demonstrate that microwear has the potential to be a powerful tool in the analysis of fish trophic ecology, particularly in the analysis of species pairs and niche differentiation. It has advantages over the trophic snapshot provided by analysis of stomach contents in that microwear reflects feeding and food preferences over a longer period of time, and can be applied where these data are unavailable. Furthermore, it is applicable to extinct organisms and fossils, allowing the role of trophic ecology, niche partitioning and competition over evolutionary time-scales to be investigated for the first time.