Genetic bias in repeated evolution of pigment loss in cave populations of the Asellus aquaticus species complex.

Similar phenotypes can evolve repeatedly under the same evolutionary pressures. A compelling example is the evolution of pigment loss and eye loss in cave-dwelling animals. While specific genomic regions or genes associated with these phenotypes have been identified in model species, it remains uncertain whether a bias towards particular genetic mechanisms exists. An isopod crustacean, Asellus aquaticus, is an ideal model organism to investigate this phenomenon. It inhabits surface freshwaters throughout Europe but has colonized groundwater on multiple independent occasions and evolved several cave populations with distinct ecomorphology. Previous studies have demonstrated that three different cave populations utilized common genetic regions, potentially the same genes, in the evolution of pigment and eye loss. Expanding on this, we conducted analysis on two additional cave populations, distinct either phylogenetically or biogeographically from those previously examined. We generated F2 hybrids from cave × surface crosses and tested phenotype-genotype associations, as well as conducted complementation tests by crossing individuals from different cave populations. Our findings revealed that pigment loss and orange eye pigment in additional cave populations were associated with the same genomic regions as observed in the three previously tested cave populations. Moreover, the lack of complementation across all cross combinations suggests that the same gene likely drives pigment loss. These results substantiate a genetic bias in the recurrent evolution of pigment loss in this model system. Future investigations should focus on the cause behind this bias, possibly arising from allele recruitment from ancestral surface populations' genetic variation or advantageous allele effects via pleiotropy.

Sexual dimorphism in subterranean amphipod crustaceans covaries with subterranean habitat type.

Sexual dimorphism can evolve in response to sex-specific selection pressures that vary across habitats. We studied sexual differences in subterranean amphipods Niphargus living in shallow subterranean habitats (close to the surface), cave streams (intermediate), and cave lakes (deepest and most isolated). These three habitats differ because at greater depths there is lower food availability, reduced predation, and weaker seasonality. Additionally, species near the surface have a near-even adult sex ratio (ASR), whereas species from cave lakes have a female-biased ASR. We hypothesized (a) a decrease in sexual dimorphism from shallow subterranean habitats to cave lake species because of weaker sexual selection derived from changes in the ASR and (b) an increase in female body size in cave lakes because of stronger fecundity selection on account of oligotrophy, reduced predation, and weaker seasonality. We measured body size and two sexually dimorphic abdominal appendages for all 31 species and several behaviours related to male competition (activity, risk-taking, exploration) for 12 species. Species with an equal ASR that live close to the surface exhibited sexual dimorphism in all three morphological traits, but not in behaviour. The body size of females increased from the surface to cave lakes, but no such trend was observed in males. In cave lake species, males and females differed neither morphologically nor behaviourally. Our results are consistent with the possibility that sexual and fecundity selection covary across the three habitats, which indirectly and directly, respectively, shape the degree of sexual dimorphism in Niphargus species.

Standing genetic variation as a potential mechanism of novel cave phenotype evolution in the freshwater isopod, Asellus aquaticus.

Novel phenotypes can come about through a variety of mechanisms including standing genetic variation from a founding population. Cave animals are an excellent system in which to study the evolution of novel phenotypes such as loss of pigmentation and eyes. Asellus aquaticus is a freshwater isopod crustacean found in Europe and has both a surface and a cave ecomorph which vary in multiple phenotypic traits. An orange eye phenotype was previously revealed by F2 crosses and backcrosses to the cave parent within two examined Slovenian cave populations. Complete loss of pigmentation, both in eye and body, is epistatic to the orange eye phenotype and therefore the orange eye phenotype is hidden within the cave populations. Our goal was to investigate the origin of the orange eye alleles within the Slovenian cave populations by examining A. aquaticus individuals from Slovenian and Romanian surface populations and Asellus aquaticus infernus individuals from a Romanian cave population. We found orange eye individuals present in lab raised surface populations of A. aquaticus from both Slovenia and Romania. Using a mapping approach with crosses between individuals of two surface populations, we found that the region known to be responsible for the orange eye phenotype within the two previously examined Slovenian cave populations was also responsible within both the Slovenian and the Romanian surface populations. Complementation crosses between orange eye Slovenian and orange eye Romanian surface individuals suggest that the same gene is responsible for the orange eye phenotype in both surface populations. Additionally, we observed a low frequency phenotype of eye loss in crosses generated between the two surface populations and also in the Romanian surface population. Finally, in a cave population from Romania, A. aquaticus infernus, we found that the same region is also responsible for the orange eye phenotype as the Slovenian cave populations and the Slovenian and Romanian surface populations. Therefore, we present evidence that variation present in the cave populations could originate from standing variation present in the surface populations and/or transgressive hybridization of different surface phylogenetic lineages rather than de novo mutations.

Screening of NaCl salinity sensitivity across eight species of subterranean amphipod genus Niphargus.

Secondary salinization of freshwater is becoming a growing environmental problem. Currently, there is few data available on the effects of salinisation on subterranean crustaceans that are vital for the maintenance of groundwater ecosystem functioning. In this study, the sensitivity of subterranean Niphargus amphipods to NaCl was investigated. We expected that cave-dwelling species would be more sensitive as surface-subterranean boundary species. Eight ecologically different Niphargus species were tested: four live at the boundary between the surface and subterranean ecosystems (N. timavi, N. krameri, N. sphagnicolus, N. spinulifemur), three live in cave streams (N. stygius, N. scopicauda, N. podpecanus), and one species (N. hebereri) lives in anchialine caves and wells. The organisms were exposed to five concentrations of NaCl for 96 h and afterwards the immobility, mortality, and electron transfer system (ETS) activity (a measure for metabolic rate of animals) were evaluated. As expected, the most tolerant species was N. hebereri dwelling in naturally high-salinity habitat. However, contrary to our expectations, the species collected at the surface-subterranean boundary were more sensitive as cave stream species when their immobility and mortality were assessed. Interestingly, the majority of Niphargus tested were more NaCl tolerant as can be deduced from currently available data for subterranean and surface crustaceans. We could not observe a clear trend in ETS activity changes between groups of surface-subterranean boundary and cave streams species after exposure to NaCl stress, but it appears that osmotic stress-induced metabolic rate changes are species-specific. This study shows that amphipods Niphargus can be a valuable subterranean environmental research model and further ecotoxicity research is of interest.

No room for males in caves: Female-biased sex ratio in subterranean amphipods of the genus Niphargus.

Sex allocation theory predicts that the proportion of daughters to sons will evolve in response to ecological conditions that determine the costs and benefits of producing each sex. All else being equal, the adult sex ratio (ASR) should also vary with ecological conditions. Many studies of subterranean species reported female-biased ASR, but no systematic study has yet been conducted. We test the hypothesis that the ASR becomes more female-biased with increased isolation from the surface. We compiled a data set of ASRs of 35 species in the subterranean amphipod Niphargus, each living in one of three distinct habitats (surface-subterranean boundary, cave streams, phreatic lakes) representing an environmental gradient of increased isolation underground. The ASR was female-biased in 27 of 35 species; the bias was statistically significant in 12 species. We found a significant difference in the ASR among habitats after correction for phylogeny. It is most weakly female-biased at the surface-subterranean boundary and most strongly female-biased in phreatic lakes. Additional modelling suggests that the ASR has evolved towards a single value for both surface-subterranean boundary and cave stream-dwelling species, and another value for 9 of 11 phreatic lake dwellers. We suggest that a history of inbreeding in subterranean populations might lower inbreeding depression such that kin selection favours mating with siblings. This could select for a female-biased offspring sex ratio due to local mate competition among brothers. The observed patterns in sex ratios in subterranean species make them a group worthy of more attention from those interested in sex allocation theory.

Light triggers habitat choice of eyeless subterranean but not of eyed surface amphipods.

Boundaries of species distributions are the result of colonization-extinction processes. Survival on the boundary depends on how well individuals discriminate optimal from suboptimal habitat patches. Such behaviour is called habitat choice and was only rarely applied to macroecology, although it links species ecological niche and species distribution. Surface and subterranean aquatic species are spatially strongly segregated, even in the absence of physical barriers. We explored whether a behavioural response to light functions as a habitat choice mechanism that could explain species turnover between surface and subterranean aquatic ecosystems. In a controlled laboratory experiment, we studied the behavioural response to light of ten pairs of surface and subterranean amphipods that permanently co-occur in springs. Surface species showed a weak photophobic, photoneutral, and in one case, photophilic response, whereas all subterranean species showed a strong photophobic response. Eyeless subterranean but not eyed surface amphipods appear to orient themselves with light cues. On a local scale, this difference possibly diminishes harmful interactions between the co-occurring amphipods, whereas on a regional scale, photophobia could explain limited dispersal and a high degree of endemism observed among subterranean species.

From darkness to twilight: Morphological divergence between cave and surface-subterranean ecotone Niphargus species.

Subterranean and surface habitats are in stark contrast in several environmental factors. Therefore, adaptation to the subterranean environment typically impedes the (re)colonisation of surface habitats. The genus Niphargus includes amphipod crustaceans that primarily occupy subterranean habitats. All its species show typical adaptations to the subterranean environment. However, some Niphargus species occur in surface-subterranean ecotones. To understand whether (i) habitat-based phenotypic divergence is present between the cave and the ecotone species and (ii) similar phenotypes emerge independently in each ecotone, we studied morphological divergence between four cave and four ecotone Niphargus species based on 13 functional morphological traits. To account for different selection acting on the sexes, we included both males (N = 244) and females (N = 222). Nine out of 13 traits showed habitat-divergence. Traits related to feeding and crawling were shorter, while traits related to oxygenation were larger in ecotone species. Eleven out of 13 traits were sexually dimorphic. Traits related to oxygenation and crawling were larger in females, while the trait related to swimming was larger in males. We found that the extent of sexual dimorphism differs between the habitats in eight traits related to sensing, feeding, oxygenation and crawling. Additionally, we found that in certain traits related to sensing and oxygenation, habitat-related differences are only present in one sex, but not the other. We conclude that the detected differences between the cave and the ecotone species indicate divergent evolution, where similarities among the different species within habitat type indicate convergent evolution. The high degree of sexual dimorphism paired with differences in sexual dimorphism between the habitats in certain traits suggest that sexual and fecundity selections have comparable effects to environmental selection. Thus, studies of habitat-dependent adaptations investigating one sex only, or not considering sexual dimorphism, can lead to erroneous conclusions.

Sex-dependent increase of movement activity in the freshwater isopod Asellus aquaticus following adaptation to a predator-free cave habitat.

Populations experiencing negligible predation pressure are expected to evolve higher behavioral activity. However, when sexes have different expected benefits from high activity, the adaptive shift is expected to be sex-specific. Here, we compared movement activity of one cave (lack of predation) and three adjacent surface (high and diverse predation) populations of Asellus aquaticus, a freshwater isopod known for its independent colonization of several caves across Europe. We predicted 1) higher activity in cave than in surface populations, with 2) the difference being more pronounced in males as they are known for active mate searching behavior, while females are not. Activity was assessed both in the presence and absence of light. Our results supported both predictions: movement activity was higher in the cave than in the surface populations, particularly in males. Relaxed predation pressure in the cave-adapted population is most likely the main selective factor behind increased behavioral activity, but we also showed that the extent of increase is sex-specific.

Avoidance behaviour toxicity tests should account for animal gregariousness: a case study on the terrestrial isopod Porcellioscaber.

Avoidance behaviour enables woodlice to escape suboptimal environmental conditions and to mitigate harmful effects of pollutants. However, several studies have shown that at least in some woodlice species the tendency to aggregate can lead to suboptimal responses as the between-conspecific attraction can outweigh the aversive stimuli. The present study evaluated the influence of gregariousness on the behaviour of Porcellioscaber in a heterogeneously polluted environment. The hypothesis was that the tendency for aggregation outweighs the tendency for exploratory activity, therefore animals in groups will be less active. Consequently, this will affect their avoidance of polluted environmental patches. To test this hypothesis, isolated individuals or pairs of individuals were monitored in free-choice arenas where animals could select between uncontaminated and pyrethrin-contaminated soils. Animals were video recorded for 3 h in darkness using infrared light and analysed for avoidance behaviour and locomotor activity. In general, isolated animals were more locomotory active and avoided the contaminated soil more than paired animals. It can be concluded that aggregation behaviour suppresses exploratory behaviour and consequently also the avoidance of polluted environments. This should be accounted for when interpreting results of avoidance tests with groups of gregarious animals, which may underestimate the effect of pollutants.

Parallel morphological evolution and habitat-dependent sexual dimorphism in cave- vs. surface populations of the Asellus aquaticus (Crustacea: Isopoda: Asellidae) species complex.

Studying parallel evolution (repeated, independent evolution of similar phenotypes in similar environments) is a powerful tool to understand environment-dependent selective forces. Surface-dwelling species that repeatedly and independently colonized caves provide unique models for such studies. The primarily surface-dwelling Asellus aquaticus species complex is a good candidate to carry out such research, because it colonized several caves in Europe. By comparing 17 functional morphological traits between six cave and nine surface populations of the A. aquaticus species complex, we investigated population divergence in morphology and sexual dimorphism. We found habitat-dependent population divergence in 10 out of 17 traits, likely reflecting habitat-driven changes in selection acting on sensory systems, feeding, grooming, and antipredator mechanisms. Sexual dimorphism was present in 15 traits, explained by sexual selection acting on male traits important in male-male agonistic behavior or mate guarding and fecundity selection acting on female traits affecting offspring number and nursing. In eight traits, the degree of sexual dimorphism was habitat dependent. We conclude that cave-related morphological changes are highly trait- and function-specific and that the strength of sexual/fecundity selection strongly differs between cave and surface habitats. The considerable population variation within habitat type warrants further studies to reveal cave-specific adaptations besides the parallel patterns.

Is foraging innovation lost following colonization of a less variable environment? A case study in surface- vs. cave-dwelling Asellus aquaticus.

Behavioral innovation is a key process for successful colonization of new habitat types. However, it is costly due to the necessary cognitive and neural demands and typically connected to ecological generalism. Therefore, loss of behavioral innovativeness is predicted following colonization of new, simple, and invariable environments. We tested this prediction by studying foraging innovativeness in the freshwater isopod Asellus aquaticus. We sampled its populations along the route of colonizing a thermokarstic water-filled cave (simple, stable habitat with only bacterial mats as food) from surface habitats (variable environment, wide variety of food). The studied cave population separated from the surface populations at least 60,000 years ago. Animals were tested both with familiar and novel food types (cave food: bacterial mats; surface food: decaying leaves). Irrespective of food type, cave individuals were more likely to feed than surface individuals. Further, animals from all populations fed longer on leaves than on bacteria, even though leaves were novel for the cave animals. Our results support that cave A. aquaticus did not lose the ability to use the ancestral (surface) food type after adapting to a simple, stable, and highly specialized habitat.

The interplay between habitat use, morphology and locomotion in subterranean crustaceans of the genus Niphargus.

Locomotion is an important, fitness-related functional trait. Environment selects for type of locomotion and shapes the morphology of locomotion-related traits such as body size and appendages. In subterranean aquatic arthropods, these traits are subjected to multiple, at times opposing selection pressures. Darkness selects for enhanced mechano- and chemosensory systems and hence elongation of appendages. Conversely, water currents have been shown to favor short appendages. However, no study has addressed the variation in locomotion of invertebrates inhabiting cave streams and cave lakes, or questioned the relationship between species' morphology and locomotion. To fill this knowledge gap, we studied the interplay between habitat use, morphology and locomotion in amphipods of the subterranean genus Niphargus. Previous studies showed that lake and stream species differ in morphology. Namely, lake species are large, stout and long-legged, whereas stream species are small, slender and short-legged. We here compared locomotion mode and speed between three lake and five stream species. In addition, we tested whether morphology predicts locomotion. We found that the stream species lie on their body sides and move using slow crawling or tail-flipping. The species inhabiting lakes move comparably faster, and use a variety of locomotion modes. Noteworthy, one of the lake species almost exclusively moves in an upright or semi-upright position that resembles walking. Body size and relative length of appendages predict locomotion mode and speed in all species. We propose that integrating locomotion in the studies of subterranean species might improve our understanding of their morphological evolution.

Fundamental research questions in subterranean biology.

Five decades ago, a landmark paper in Science titled The Cave Environment heralded caves as ideal natural experimental laboratories in which to develop and address general questions in geology, ecology, biogeography, and evolutionary biology. Although the 'caves as laboratory' paradigm has since been advocated by subterranean biologists, there are few examples of studies that successfully translated their results into general principles. The contemporary era of big data, modelling tools, and revolutionary advances in genetics and (meta)genomics provides an opportunity to revisit unresolved questions and challenges, as well as examine promising new avenues of research in subterranean biology. Accordingly, we have developed a roadmap to guide future research endeavours in subterranean biology by adapting a well-established methodology of 'horizon scanning' to identify the highest priority research questions across six subject areas. Based on the expert opinion of 30 scientists from around the globe with complementary expertise and of different academic ages, we assembled an initial list of 258 fundamental questions concentrating on macroecology and microbial ecology, adaptation, evolution, and conservation. Subsequently, through online surveys, 130 subterranean biologists with various backgrounds assisted us in reducing our list to 50 top-priority questions. These research questions are broad in scope and ready to be addressed in the next decade. We believe this exercise will stimulate research towards a deeper understanding of subterranean biology and foster hypothesis-driven studies likely to resonate broadly from the traditional boundaries of this field.

No need to hide in caves: shelter-seeking behavior of surface and cave ecomorphs of Asellus aquaticus (Isopoda: Crustacea).

Shelter-seeking is a vital behavior for stress reduction and survival in a range of animals. It comes at the cost of reduced foraging, mate finding, dispersal and territoriality, and is expected to reflect the trade-off between fitness costs and benefits. One way to test this hypothesis is to compare shelter-seeking behavior in surface habitats and in caves where external threat factors are largely reduced. We did so using the freshwater isopod Asellus aquaticus from the Postojna-Planina Cave System and surrounding surface waters. Animals from two distinct, replicated pairs of surface and cave populations were individually offered a choice between shelter and open area. The thigmotatic sensation of a transparent plastic plate was the only stimulus that could trigger the sheltering behavior. Video recordings showed a clear reduction of shelter-seeking behavior in the cave ecomorph in one population pair (Pivka). There were no changes in the other population pair (Rak), where the behavioral response had a much higher variance. Our results were partly in agreement with the hypothesis that shelter-seeking behavior should be selected against in an environment with reduced external threats. It is nevertheless too early for generalizations as the results in the second population pair were inconclusive. Additionally, we showed that for benthic walkers like A. aquaticus the use of rough substrate is crucial to obtain unbiased behavioral responses. Results of some previous studies using smooth glass or plastic substratum could be affected by unnatural behavior of animals constantly trying to find firm contact with the ground.

Description of Acanthocephalus anguillae balkanicus subsp. n. (Acanthocephala: Echinorhynchidae) from Proteus anguinus Laurenti (Amphibia: Proteidae) and the cave ecomorph of Asellus aquaticus (Crustacea: Asellidae) in Slovenia.

Acanthocephalus balkanicus Batchvarov et Combes, 1974 was incompletely described from the northern crested newt, Triturus cristatus (Laurenti) (Amphibia: Salamandridae), a possible synonym of the Balkan crested newt, Triturus ivanbureschi Arntzen et Wielstra, from a pond in village of Pesnopoy, southern Bulgaria. We provide a full description of adult males and females of the same taxon from the olm, Proteus anguinus Laurenti (Amphibia: Proteidae), the only exclusively aquatic cave-dwelling vertebrate in Europe, captured in Postojna-Planina Cave System in Slovenia. Cystacanths were also collected from the cave ecomorph of Asellus aquaticus (Linnaeus) (Crustacea: Asellidae) in the same location. Molecular analysis of specimens from Slovenia revealed that they are genetically almost identical to those of Acanthocephalus anguillae (Müller, 1780), a common parasite of European freshwater fishes. We propose to recognise the morphological and host differences by describing A. balkanicus as a new subspecies of A. anguillae. Acanthocephalus anguillae balkanicus is rather small and cylindrical with cylindrical proboscis having 10 rows of 6 hooks with simple roots each, long neck, large balloon-shaped lemnisci, small spherical anterior testis, and 6 club-shaped cement glands in 3 pairs. SEM images reveal more morphological details and the X-ray scans of gallium cut hooks shows considerably higher levels of phosphorus and calcium in adult hooks than in cystacanth hooks, especially in basal areas. Sulfur levels were higher in the arch and basal area of cystacanth hooks than adult hooks. Considering that both definitive and intermediate hosts of the Slovenian population of this acanthocephalan are bound to cave life, it is possible that its entire life cycle is uniquely completed underground.

Common Genetic Basis of Eye and Pigment Loss in Two Distinct Cave Populations of the Isopod Crustacean Asellus aquaticus.

Repeated evolution of similar phenotypes is a widespread phenomenon found throughout the living world and it can proceed through the same or different genetic mechanisms. Cave animals with their convergent traits such as eye and pigment loss, as well as elongated appendages, are a striking example of the evolution of similar phenotypes. Yet, few cave species are amenable to genetic crossing and mapping techniques making it challenging to determine the genetic mechanisms causing their similar phenotypes. To address this limitation, we have been developing Asellus aquaticus, a freshwater isopod crustacean, as a genetic model. Many of its cave populations originate from separate colonization events and thus independently evolved their similar cave-related phenotypes which differ from the still existent ancestral-like surface populations. In our prior work, we identified genomic regions responsible for eye and pigment loss in a single cave population from Slovenia. In this study we examined another, independently evolved cave population, also from Slovenia, and asked whether the same or different genomic regions are responsible for eye and pigment loss in the two cave populations. We generated F2 and backcross hybrids with a surface population, genotyped them for the previously identified genomic regions, and performed a complementation test by crossing individuals from the two cave populations. We found out that the same genomic regions are responsible for eye and pigment loss and that at least one of the genes causing pigment loss is the same in both cave populations. Future studies will identify the actual genes and mutations, as well as examine additional cave populations to see if the same genes are commonly associated with eye and pigment loss in this species.

Comparative study of acetylcholinesterase and glutathione S-transferase activities of closely related cave and surface Asellus aquaticus (Isopoda: Crustacea).

The freshwater isopod crustacean Asellus aquaticus has recently been developed as an emerging invertebrate cave model for studying evolutionary and developmental biology. Mostly morphological and genetic differences between cave and surface A. aquaticus populations have been described up to now, while scarce data are available on other aspects, including physiology. The purpose of this study was to advance our understanding of the physiological differences between cave A. aquaticus and its surface-dwelling counterparts. We sampled two surface populations from the surface section of the sinking Pivka River (central Slovenia, Europe), i.e. locality Pivka Polje, and locality Planina Polje, and one cave population from the subterranean section of the sinking Pivka River, i.e. locality Planina Cave. Animals were sampled in spring, summer and autumn. We measured the activities of acetylcholinesterase (AChE) and glutathione S-transferase (GST) in individuals snap-frozen in the field immediately after collection. Acetylcholinesterase is likely related to animals' locomotor activity, while GST activity is related to the metabolic activity of an organism. Our study shows significantly lower AChE and GST activities in the cave population in comparison to both surface A. aquaticus populations. This confirms the assumption that cave A. aquaticus have lower locomotor and metabolic activity than surface A. aquaticus in their respective natural environments. In surface A. aquaticus populations, seasonal fluctuations in GST activity were observed, while these were less pronounced in individuals from the more stable cave environment. On the other hand, AChE activity was generally season-independent in all populations. To our knowledge, this is the first study of its kind conducted in A. aquaticus. Our results show that among closely related cave and surface A. aquaticus populations also physiological differences are present besides the morphological and genetic. These findings contribute to a better understanding of the biology of A. aquaticus and cave crustaceans in general.

Morphologically Cryptic Amphipod Species Are "Ecological Clones" at Regional but Not at Local Scale: A Case Study of Four Niphargus Species.

Recent studies indicate that morphologically cryptic species may be ecologically more different than would be predicted from their morphological similarity and phylogenetic relatedness. However, in biodiversity research it often remains unclear whether cryptic species should be treated as ecologically equivalent, or whether detected differences have ecological significance. In this study, we assessed the ecological equivalence of four morphologically cryptic species of the amphipod genus Niphargus. All species live in a small, isolated area on the Istrian Peninsula in the NW Balkans. The distributional ranges of the species are partially overlapping and all species are living in springs. We reconstructed their ecological niches using morphological traits related to feeding, bioclimatic niche envelope and species' preference for epi-hypogean habitats. The ecological meaning of differences in niches was evaluated using distributional data and co-occurrence frequencies. We show that the species comprise two pairs of sister species. All species differ from each other and the degree of differentiation is not related to phylogenetic relatedness. Moreover, low co-occurrence frequencies in sympatric zones imply present or past interspecific competition. This pattern suggests that species are not differentiated enough to reduce interspecific competition, nor ecologically equivalent to co-exist via neutral dynamics. We tentatively conclude that the question of ecological equivalence relates to the scale of the study: at a fine scale, species' differences may influence dynamics in a local community, whereas at the regional level these species likely play roughly similar ecological roles.