Grazing herbivores reduce herbaceous biomass and fire activity across African savannas.

Fire and herbivory interact to alter ecosystems and carbon cycling. In savannas, herbivores can reduce fire activity by removing grass biomass, but the size of these effects and what regulates them remain uncertain. To examine grazing effects on fuels and fire regimes across African savannas, we combined data from herbivore exclosure experiments with remotely sensed data on fire activity and herbivore density. We show that, broadly across African savannas, grazing herbivores substantially reduce both herbaceous biomass and fire activity. The size of these effects was strongly associated with grazing herbivore densities, and surprisingly, was mostly consistent across different environments. A one-zebra increase in herbivore biomass density (~100 kg/km2 of metabolic biomass) resulted in a ~53 kg/ha reduction in standing herbaceous biomass and a ~0.43 percentage point reduction in burned area. Our results indicate that fire models can be improved by incorporating grazing effects on grass biomass.

A CRISPR-based strategy for targeted sequencing in biodiversity science.

Many applications in molecular ecology require the ability to match specific DNA sequences from single- or mixed-species samples with a diagnostic reference library. Widely used methods for DNA barcoding and metabarcoding employ PCR and amplicon sequencing to identify taxa based on target sequences, but the target-specific enrichment capabilities of CRISPR-Cas systems may offer advantages in some applications. We identified 54,837 CRISPR-Cas guide RNAs that may be useful for enriching chloroplast DNA across phylogenetically diverse plant species. We tested a subset of 17 guide RNAs in vitro to enrich plant DNA strands ranging in size from diagnostic DNA barcodes of 1,428 bp to entire chloroplast genomes of 121,284 bp. We used an Oxford Nanopore sequencer to evaluate sequencing success based on both single- and mixed-species samples, which yielded mean chloroplast sequence lengths of 2,530-11,367 bp, depending on the experiment. In comparison to mixed-species experiments, single-species experiments yielded more on-target sequence reads and greater mean pairwise identity between contigs and the plant species' reference genomes. But nevertheless, these mixed-species experiments yielded sufficient data to provide ≥48-fold increase in sequence length and better estimates of relative abundance for a commercially prepared mixture of plant species compared to DNA metabarcoding based on the chloroplast trnL-P6 marker. Prior work developed CRISPR-based enrichment protocols for long-read sequencing and our experiments pioneered its use for plant DNA barcoding and chloroplast assemblies that may have advantages over workflows that require PCR and short-read sequencing. Future work would benefit from continuing to develop in vitro and in silico methods for CRISPR-based analyses of mixed-species samples, especially when the appropriate reference genomes for contig assembly cannot be known a priori.

Foraging history of individual elephants using DNA metabarcoding.

Individual animals should adjust diets according to food availability. We used DNA metabarcoding to construct individual-level dietary timeseries for elephants from two family groups in Kenya varying in habitat use, social position and reproductive status. We detected at least 367 dietary plant taxa, with up to 137 unique plant sequences in one fecal sample. Results matched well-established trends: elephants tended to eat more grass when it rained and other plants when dry. Nested within these switches from 'grazing' to 'browsing' strategies, dietary DNA revealed seasonal shifts in food richness, composition and overlap between individuals. Elephants of both families converged on relatively cohesive diets in dry seasons but varied in their maintenance of cohesion during wet seasons. Dietary cohesion throughout the timeseries of the subdominant 'Artists' family was stronger and more consistently positive compared to the dominant 'Royals' family. The greater degree of individuality within the dominant family's timeseries could reflect more divergent nutritional requirements associated with calf dependency and/or priority access to preferred habitats. Whereas theory predicts that individuals should specialize on different foods under resource scarcity, our data suggest family bonds may promote cohesion and foster the emergence of diverse feeding cultures reflecting links between social behaviour and nutrition.

The Difficulty of Predicting Evolutionary Change in Response to Novel Ecological Interactions: A Field Experiment with Anolis Lizards.

AbstractDetermining whether and how evolution is predictable is an important goal, particularly as anthropogenic disturbances lead to novel species interactions that could modify selective pressures. Here, we use a multigeneration field experiment with brown anole lizards (Anolis sagrei) to test hypotheses about the predictability of evolution. We manipulated the presence/absence of predators and competitors of A. sagrei across 16 islands in the Bahamas that had preexisting brown anole populations. Before the experiment and again after roughly five generations, we measured traits related to locomotor performance and habitat use by brown anoles and used double-digest restriction enzyme-associated DNA sequencing to estimate genome-wide changes in allele frequencies. Although previous work showed that predators and competitors had characteristic effects on brown anole behavior, diet, and population sizes, we found that evolutionary change at both phenotypic and genomic levels was difficult to forecast. Phenotypic changes were contingent on sex and habitat use, whereas genetic change was unpredictable and not measurably correlated with phenotypic changes, experimental treatments, or other environmental factors. Our work shows how differences in ecological context can alter evolutionary outcomes over short timescales and underscores the difficulty of forecasting evolutionary responses to multispecies interactions in natural conditions, even in a well-studied system with ample supporting ecological information.

Host phylogeny and functional traits differentiate gut microbiomes in a diverse natural community of small mammals.

Differences in the bacterial communities inhabiting mammalian gut microbiomes tend to reflect the phylogenetic relatedness of their hosts, a pattern dubbed phylosymbiosis. Although most research on this pattern has compared the gut microbiomes of host species across biomes, understanding the evolutionary and ecological processes that generate phylosymbiosis requires comparisons across phylogenetic scales and under similar ecological conditions. We analysed the gut microbiomes of 14 sympatric small mammal species in a semi-arid African savanna, hypothesizing that there would be a strong phylosymbiotic pattern associated with differences in their body sizes and diets. Consistent with phylosymbiosis, microbiome dissimilarity increased with phylogenetic distance among hosts, ranging from congeneric sets of mice and hares that did not differ significantly in microbiome composition to species from different taxonomic orders that had almost no gut bacteria in common. While phylosymbiosis was detected among just the 11 species of rodents, it was substantially weaker at this scale than in comparisons involving all 14 species together. In contrast, microbiome diversity and composition were generally more strongly correlated with body size, dietary breadth, and dietary overlap in comparisons restricted to rodents than in those including all lineages. The starkest divides in microbiome composition thus reflected the broad evolutionary divergence of hosts, regardless of body size or diet, while subtler microbiome differences reflected variation in ecologically important traits of closely related hosts. Strong phylosymbiotic patterns arose deep in the phylogeny, and ecological filters that promote functional differentiation of cooccurring host species may disrupt or obscure this pattern near the tips.

Evidence-based strategies to navigate complexity in dietary DNA metabarcoding: A reply.

It is clearly beneficial to eliminate low-abundance sequences that arise in error during dietary DNA metabarcoding studies, but to purge all low-abundance sequences is to risk eliminating real sequences and complicating ecological analyses. Our prior literature review noted that DNA sequence relative read abundance (RRA) thresholds can help ameliorate false-positive taxon occurrences, but that historical emphasis on this utility has fostered uncertainty about the associated risk of inflating the false-negative rate (Littleford-Colquhoun et al., 2022). To address this, we combined a simulation study and an empirical data set to both illustrate the issue and provide blueprints for simulation studies and sensitivity analyses that can help investigators avoid overcorrecting and thereby bolster confidence in ecological inferences. Awareness of both the costs and the benefits of abundance-filtering is needed because accurately characterizing dietary distributions can be critically important for understanding animal diets, nutrition and trophic networks. Highlighting the need to raise awareness, a critique of our paper emphasized the misleading notion that "false positive interactions between species can present fundamentally incorrect network structures in network ecology, whereas false negatives will provide a correct but incomplete version of the network" (Tercel & Cuff, 2022). Asserting that the reliability of results will be eroded by false positives but resilient to the omission of true positives is risky and runs counter to evidence. Unfortunately, abundance-filtering methods can introduce false negatives at higher rates than they eliminate false positives and thereby undermine the analysis of otherwise reliable sequencing data. Overcorrecting can qualitatively alter and ultimately undermine ecological interpretations.

The generality of cryptic dietary niche differences in diverse large-herbivore assemblages.

Ecological niche differences are necessary for stable species coexistence but are often difficult to discern. Models of dietary niche differentiation in large mammalian herbivores invoke the quality, quantity, and spatiotemporal distribution of plant tissues and growth forms but are agnostic toward food plant species identity. Empirical support for these models is variable, suggesting that additional mechanisms of resource partitioning may be important in sustaining large-herbivore diversity in African savannas. We used DNA metabarcoding to conduct a taxonomically explicit analysis of large-herbivore diets across southeastern Africa, analyzing ∼4,000 fecal samples of 30 species from 10 sites in seven countries over 6 y. We detected 893 food plant taxa from 124 families, but just two families-grasses and legumes-accounted for the majority of herbivore diets. Nonetheless, herbivore species almost invariably partitioned food plant taxa; diet composition differed significantly in 97% of pairwise comparisons between sympatric species, and dissimilarity was pronounced even between the strictest grazers (grass eaters), strictest browsers (nongrass eaters), and closest relatives at each site. Niche differentiation was weakest in an ecosystem recovering from catastrophic defaunation, indicating that food plant partitioning is driven by species interactions, and was stronger at low rainfall, as expected if interspecific competition is a predominant driver. Diets differed more between browsers than grazers, which predictably shaped community organization: Grazer-dominated trophic networks had higher nestedness and lower modularity. That dietary differentiation is structured along taxonomic lines complements prior work on how herbivores partition plant parts and patches and suggests that common mechanisms govern herbivore coexistence and community assembly in savannas.

Large-herbivore nemabiomes: patterns of parasite diversity and sharing.

Amidst global shifts in the distribution and abundance of wildlife and livestock, we have only a rudimentary understanding of ungulate parasite communities and parasite-sharing patterns. We used qPCR and DNA metabarcoding of fecal samples to characterize gastrointestinal nematode (Strongylida) community composition and sharing among 17 sympatric species of wild and domestic large mammalian herbivore in central Kenya. We tested a suite of hypothesis-driven predictions about the role of host traits and phylogenetic relatedness in describing parasite infections. Host species identity explained 27-53% of individual variation in parasite prevalence, richness, community composition and phylogenetic diversity. Host and parasite phylogenies were congruent, host gut morphology predicted parasite community composition and prevalence, and hosts with low evolutionary distinctiveness were centrally positioned in the parasite-sharing network. We found no evidence that host body size, social-group size or feeding height were correlated with parasite composition. Our results highlight the interwoven evolutionary and ecological histories of large herbivores and their gastrointestinal nematodes and suggest that host identity, phylogeny and gut architecture-a phylogenetically conserved trait related to parasite habitat-are the overriding influences on parasite communities. These findings have implications for wildlife management and conservation as wild herbivores are increasingly replaced by livestock.

The precautionary principle and dietary DNA metabarcoding: Commonly used abundance thresholds change ecological interpretation.

Dietary DNA metabarcoding enables researchers to identify and characterize trophic interactions with a high degree of taxonomic precision. It is also sensitive to sources of bias and contamination in the field and laboratory. One of the earliest and most common strategies for dealing with such sensitivities has been to remove all low-abundance sequences and conduct ecological analyses based on the presence or absence of food taxa. Although this step is now often perceived to be necessary, evidence of its sufficiency is lacking and more attention to the risk of introducing other errors is needed. Using computer simulations, we demonstrate that common strategies to remove low-abundance sequences can erroneously eliminate true dietary sequences in ways that impact downstream inferences. Using real data from well-studied wildlife populations in Yellowstone National Park, we further show how these strategies can markedly alter the composition of dietary profiles in ways that scale-up to obscure ecological interpretations about dietary generalism, specialism, and composition. Although the practice of removing low-abundance sequences may continue to be a useful strategy to address research questions that focus on a subset of relatively abundant foods, its continued widespread use risks generating misleading perceptions about the structure of trophic networks. Researchers working with dietary DNA metabarcoding data-or similar data such as environmental DNA, microbiomes, or pathobiomes-should be aware of drawbacks and consider alternative bioinformatic, experimental, and statistical solutions.

Ecological consequences of large herbivore exclusion in an African savanna: 12 years of data from the UHURU experiment.

Diverse communities of large mammalian herbivores (LMH), once widespread, are now rare. LMH exert strong direct and indirect effects on community structure and ecosystem functions, and measuring these effects is important for testing ecological theory and for understanding past, current, and future environmental change. This in turn requires long-term experimental manipulations, owing to the slow and often nonlinear responses of populations and assemblages to LMH removal. Moreover, the effects of particular species or body-size classes within diverse LMH guilds are difficult to pinpoint, and the magnitude and even direction of these effects often depends on environmental context. Since 2008, we have maintained the Ungulate Herbivory Under Rainfall Uncertainty (UHURU) experiment, a series of size-selective LMH exclosures replicated across a rainfall/productivity gradient in a semiarid Kenyan savanna. The goals of the UHURU experiment are to measure the effects of removing successively smaller size classes of LMH (mimicking the process of size-biased extirpation) and to establish how these effects are shaped by spatial and temporal variation in rainfall. The UHURU experiment comprises three LMH-exclusion treatments and an unfenced control, applied to nine randomized blocks of contiguous 1-ha plots (n = 36). The fenced treatments are MEGA (exclusion of megaherbivores, elephant and giraffe), MESO (exclusion of herbivores ≥40 kg), and TOTAL (exclusion of herbivores ≥5 kg). Each block is replicated three times at three sites across the 20-km rainfall gradient, which has fluctuated over the course of the experiment. The first 5 years of data were published previously (Ecological Archives E095-064) and have been used in numerous studies. Since that publication, we have (1) continued to collect data following the original protocols, (2) improved the taxonomic resolution and accuracy of plant and small-mammal identifications, and (3) begun collecting several new data sets. Here, we present updated and extended raw data from the first 12 years of the UHURU experiment (2008-2019). Data include daily rainfall data throughout the experiment; annual surveys of understory plant communities; annual censuses of woody-plant communities; annual measurements of individually tagged woody plants; monthly monitoring of flowering and fruiting phenology; every-other-month small-mammal mark-recapture data; and quarterly large-mammal dung surveys. There are no copyright restrictions; notification of when and how data are used is appreciated and users of UHURU data should cite this data paper when using the data.

Large herbivores suppress liana infestation in an African savanna.

African savannas are the last stronghold of diverse large-mammal communities, and a major focus of savanna ecology is to understand how these animals affect the relative abundance of trees and grasses. However, savannas support diverse plant life-forms, and human-induced changes in large-herbivore assemblages-declining wildlife populations and their displacement by livestock-may cause unexpected shifts in plant community composition. We investigated how herbivory affects the prevalence of lianas (woody vines) and their impact on trees in an East African savanna. Although scarce (<2% of tree canopy area) and defended by toxic latex, the dominant liana, Cynanchum viminale (Apocynaceae), was eaten by 15 wild large-herbivore species and was consumed in bulk by native browsers during experimental cafeteria trials. In contrast, domesticated ungulates rarely ate lianas. When we experimentally excluded all large herbivores for periods of 8 to 17 y (simulating extirpation), liana abundance increased dramatically, with up to 75% of trees infested. Piecewise exclusion of different-sized herbivores revealed functional complementarity among size classes in suppressing lianas. Liana infestation reduced tree growth and reproduction, but herbivores quickly cleared lianas from trees after the removal of 18-y-old exclosure fences (simulating rewilding). A simple model of liana contagion showed that, without herbivores, the long-term equilibrium could be either endemic (liana-tree coexistence) or an all-liana alternative stable state. We conclude that ongoing declines of wild large-herbivore populations will disrupt the structure and functioning of many African savannas in ways that have received little attention and that may not be mitigated by replacing wildlife with livestock.

Experimental evidence that effects of megaherbivores on mesoherbivore space use are influenced by species' traits.

The extinction of 80% of megaherbivore (>1,000 kg) species towards the end of the Pleistocene altered vegetation structure, fire dynamics and nutrient cycling world-wide. Ecologists have proposed (re)introducing megaherbivores or their ecological analogues to restore lost ecosystem functions and reinforce extant but declining megaherbivore populations. However, the effects of megaherbivores on smaller herbivores are poorly understood. We used long-term exclusion experiments and multispecies hierarchical models fitted to dung counts to test (a) the effect of megaherbivores (elephant and giraffe) on the occurrence (dung presence) and use intensity (dung pile density) of mesoherbivores (2-1,000 kg), and (b) the extent to which the responses of each mesoherbivore species was predictable based on their traits (diet and shoulder height) and phylogenetic relatedness. Megaherbivores increased the predicted occurrence and use intensity of zebras but reduced the occurrence and use intensity of several other mesoherbivore species. The negative effect of megaherbivores on mesoherbivore occurrence was stronger for shorter species, regardless of diet or relatedness. Megaherbivores substantially reduced the expected total use intensity (i.e. cumulative dung density of all species) of mesoherbivores, but only minimally reduced the expected species richness (i.e. cumulative predicted occurrence probabilities of all species) of mesoherbivores (by <1 species). Simulated extirpation of megaherbivores altered use intensity by mesoherbivores, which should be considered during (re)introductions of megaherbivores or their ecological proxies. Species' traits (in this case shoulder height) may be more reliable predictors of mesoherbivores' responses to megaherbivores than phylogenetic relatedness, and may be useful for predicting responses of data-limited species.

Group 2i Isochrysidales produce characteristic alkenones reflecting sea ice distribution.

Alkenones are biomarkers produced solely by algae in the order Isochrysidales that have been used to reconstruct sea surface temperature (SST) since the 1980s. However, alkenone-based SST reconstructions in the northern high latitude oceans show significant bias towards warmer temperatures in core-tops, diverge from other SST proxies in down core records, and are often accompanied by anomalously high relative abundance of the C37 tetra-unsaturated methyl alkenone (%C37:4). Elevated %C37:4 is widely interpreted as an indicator of low sea surface salinity from polar water masses, but its biological source has thus far remained elusive. Here we identify a lineage of Isochrysidales that is responsible for elevated C37:4 methyl alkenone in the northern high latitude oceans through next-generation sequencing and lab-culture experiments. This Isochrysidales lineage co-occurs widely with sea ice in marine environments and is distinct from other known marine alkenone-producers, namely Emiliania huxleyi and Gephyrocapsa oceanica. More importantly, the %C37:4 in seawater filtered particulate organic matter and surface sediments is significantly correlated with annual mean sea ice concentrations. In sediment cores from the Svalbard region, the %C37:4 concentration aligns with the Greenland temperature record and other qualitative regional sea ice records spanning the past 14 kyrs, reflecting sea ice concentrations quantitatively. Our findings imply that %C37:4 is a powerful proxy for reconstructing sea ice conditions in the high latitude oceans on thousand- and, potentially, on million-year timescales.

Multiple dimensions of dietary diversity in large mammalian herbivores.

Theory predicts that trophic specialization (i.e. low dietary diversity) should make consumer populations sensitive to environmental disturbances. Yet diagnosing specialization is complicated both by the difficulty of precisely quantifying diet composition and by definitional ambiguity: what makes a diet 'diverse'? We sought to characterize the relationship between taxonomic dietary diversity (TDD) and phylogenetic dietary diversity (PDD) in a species-rich community of large mammalian herbivores in a semi-arid East African savanna. We hypothesized that TDD and PDD would be positively correlated within and among species, because taxonomically diverse diets are likely to include plants from many lineages. By using DNA metabarcoding to analyse 1,281 faecal samples collected across multiple seasons, we compiled high-resolution diet profiles for 25 sympatric large-herbivore species. For each of these populations, we calculated TDD and PDD with reference to a DNA reference library for local plants. Contrary to our hypothesis, measures of TDD and PDD were either uncorrelated or negatively correlated with each other. Thus, these metrics reflect distinct dimensions of dietary specialization both within and among species. In general, grazers and ruminants exhibited greater TDD, but lower PDD, than did browsers and non-ruminants. We found significant seasonal variation in TDD and/or PDD for all but four species (Grevy's zebra, buffalo, elephant, Grant's gazelle); however, the relationship between TDD and PDD was consistent across seasons for all but one of the 12 best-sampled species (plains zebra). Our results show that taxonomic generalists can be phylogenetic specialists, and vice versa. These two dimensions of dietary diversity suggest contrasting implications for efforts to predict how consumers will respond to climate change and other environmental perturbations. For example, populations with low TDD may be sensitive to phylogenetically 'random' losses of food species, whereas populations with low PDD may be comparatively more sensitive to environmental changes that disadvantage entire plant lineages-and populations with low dietary diversity in both taxonomic and phylogenetic dimensions may be most vulnerable of all.

Trophic rewilding revives biotic resistance to shrub invasion.

Trophic rewilding seeks to rehabilitate degraded ecosystems by repopulating them with large animals, thereby re-establishing strong top-down interactions. Yet there are very few tests of whether such initiatives can restore ecosystem structure and functions, and on what timescales. Here we show that war-induced collapse of large-mammal populations in Mozambique's Gorongosa National Park exacerbated woody encroachment by the invasive shrub Mimosa pigra-considered one of the world's 100 worst invasive species-and that one decade of concerted trophic rewilding restored this invasion to pre-war baseline levels. Mimosa occurrence increased between 1972 and 2015, a period encompassing the near extirpation of large herbivores during the Mozambican Civil War. From 2015 to 2019, mimosa abundance declined as ungulate biomass recovered. DNA metabarcoding revealed that ruminant herbivores fed heavily on mimosa, and experimental exclosures confirmed the causal role of mammalian herbivory in containing shrub encroachment. Our results provide mechanistic evidence that trophic rewilding has rapidly revived a key ecosystem function (biotic resistance to a notorious woody invader), underscoring the potential for restoring ecological health in degraded protected areas.

Covariation of diet and gut microbiome in African megafauna.

A major challenge in biology is to understand how phylogeny, diet, and environment shape the mammalian gut microbiome. Yet most studies of nonhuman microbiomes have relied on relatively coarse dietary categorizations and have focused either on individual wild populations or on captive animals that are sheltered from environmental pressures, which may obscure the effects of dietary and environmental variation on microbiome composition in diverse natural communities. We analyzed plant and bacterial DNA in fecal samples from an assemblage of 33 sympatric large-herbivore species (27 native, 6 domesticated) in a semiarid East African savanna, which enabled high-resolution assessment of seasonal variation in both diet and microbiome composition. Phylogenetic relatedness strongly predicted microbiome composition (r = 0.91) and was weakly but significantly correlated with diet composition (r = 0.20). Dietary diversity did not significantly predict microbiome diversity across species or within any species except kudu; however, diet composition was significantly correlated with microbiome composition both across and within most species. We found a spectrum of seasonal sensitivity at the diet-microbiome nexus: Seasonal changes in diet composition explained 25% of seasonal variation in microbiome composition across species. Species' positions on (and deviations from) this spectrum were not obviously driven by phylogeny, body size, digestive strategy, or diet composition; however, domesticated species tended to exhibit greater diet-microbiome turnover than wildlife. Our results reveal marked differences in the influence of environment on the degree of diet-microbiome covariation in free-ranging African megafauna, and this variation is not well explained by canonical predictors of nutritional ecology.

Using DNA Metabarcoding To Evaluate the Plant Component of Human Diets: a Proof of Concept.

Dietary intake is difficult to measure reliably in humans because approaches typically rely on self-reporting, which can be incomplete and biased. In field studies of animals, DNA sequencing-based approaches such as metabarcoding have been developed to characterize diets, but such approaches have not previously been widely applied to humans. Here, we present data derived from sequencing of a chloroplast DNA marker (the P6 loop of the trnL [UAA] intron) in stool samples collected from 11 individuals consuming both controlled and freely selected diets. The DNA metabarcoding strategy resulted in successful PCR amplification in about 50% of samples, which increased to a 70% success rate in samples from individuals eating a controlled plant-rich diet. Detection of plant taxa among sequenced samples yielded a recall of 0.86 and a precision of 0.55 compared to a written diet record during controlled feeding of plant-based foods. The majority of sequenced plant DNA matched common human food plants, including grains, vegetables, fruits, and herbs prepared both cooked and uncooked. Moreover, DNA metabarcoding data were sufficient to distinguish between baseline and treatment diet arms of the study. Still, the relatively high PCR failure rate and an inability to distinguish some dietary plants at the sequence level using the trnL-P6 marker suggest that future methodological refinements are necessary. Overall, our results suggest that DNA metabarcoding provides a promising new method for tracking human plant intake and that similar approaches could be used to characterize the animal and fungal components of our omnivorous diets.IMPORTANCE Current methods for capturing human dietary patterns typically rely on individual recall and as such are subject to the limitations of human memory. DNA sequencing-based approaches, frequently used for profiling nonhuman diets, do not suffer from the same limitations. Here, we used metabarcoding to broadly characterize the plant portion of human diets for the first time. The majority of sequences corresponded to known human foods, including all but one foodstuff included in an experimental plant-rich diet. Metabarcoding could distinguish between experimental diets and matched individual diet records from controlled settings with high accuracy. Because this method is independent of survey language and timing, it could also be applied to geographically and culturally disparate human populations, as well as in retrospective studies involving banked human stool.

Predator-induced collapse of niche structure and species coexistence.

Biological invasions are both a pressing environmental challenge and an opportunity to investigate fundamental ecological processes, such as the role of top predators in regulating biodiversity and food-web structure. In whole-ecosystem manipulations of small Caribbean islands on which brown anole lizards (Anolis sagrei) were the native top predator, we experimentally staged invasions by competitors (green anoles, Anolis smaragdinus) and/or new top predators (curly-tailed lizards, Leiocephalus carinatus). We show that curly-tailed lizards destabilized the coexistence of competing prey species, contrary to the classic idea of keystone predation. Fear-driven avoidance of predators collapsed the spatial and dietary niche structure that otherwise stabilized coexistence, which intensified interspecific competition within predator-free refuges and contributed to the extinction of green-anole populations on two islands. Moreover, whereas adding either green anoles or curly-tailed lizards lengthened food chains on the islands, adding both species reversed this effect-in part because the apex predators were trophic omnivores. Our results underscore the importance of top-down control in ecological communities, but show that its outcomes depend on prey behaviour, spatial structure, and omnivory. Diversity-enhancing effects of top predators cannot be assumed, and non-consumptive effects of predation risk may be a widespread constraint on species coexistence.

Phylogeography of the Neotropical epiphytic orchid, Brassavola nodosa: evidence for a secondary contact zone in northwestern Costa Rica.

Spatial patterns of genetic variation can reveal otherwise cryptic evolutionary and landscape processes. In northwestern Costa Rica, an approximately concordant genetic discontinuity occurs among populations of several plant species. We conducted phylogeographic analyses of an epiphytic orchid, Brassavola nodosa, to test for genetic discontinuity and to explore its underlying causes. We genotyped 18 populations with 19 nuclear loci and two non-coding chloroplast sequence regions. We estimated genetic diversity and structure, relative importance of pollen and seed dispersal, and divergence time to understand how genetic diversity was spatially partitioned. Nuclear genetic diversity was high with little differentiation among populations (GSTn = 0.065). In contrast, chloroplast haplotypes were highly structured (GSTc = 0.570) and reveal a discontinuity between northwestern and southeastern populations within Costa Rica. Haplotype differences suggest two formerly isolated lineages that diverged ~10,000-100,000 YBP. Haplotype mixing and greater genetic diversity occur in an intermediate transition zone. Patterns of nuclear and chloroplast data were consistent. Different levels of genetic differentiation for the two genomes reflect the relative effectiveness of biotic versus abiotic dispersers of pollen and seeds, respectively. Isolation of the two lineages likely resulted from the complex environmental and geophysical history of the region. Our results suggest a recent cryptic seed dispersal barrier and/or zone of secondary contact. We hypothesize that powerful northeasterly trade winds hinder movement of wind-borne seeds between the two regions, while the multi-directional dispersal of pollen by strong-flying sphinx moths resulted in lower differentiation of nuclear loci.

Plant DNA-barcode library and community phylogeny for a semi-arid East African savanna.

Applications of DNA barcoding include identifying species, inferring ecological and evolutionary relationships between species, and DNA metabarcoding. These applications require reference libraries that are not yet available for many taxa and geographic regions. We collected, identified, and vouchered plant specimens from Mpala Research Center in Laikipia, Kenya, to develop an extensive DNA-barcode library for a savanna ecosystem in equatorial East Africa. We amassed up to five DNA barcode markers (rbcL, matK, trnL-F, trnH-psbA, and ITS) for 1,781 specimens representing up to 460 species (~92% of the known flora), increasing the number of plant DNA barcode records for Africa by ~9%. We evaluated the ability of these markers, singly and in combination, to delimit species by calculating intra- and interspecific genetic distances. We further estimated a plant community phylogeny and demonstrated its utility by testing if evolutionary relatedness could predict the tendency of members of the Mpala plant community to have or lack "barcode gaps", defined as disparities between the maximum intra- and minimum interspecific genetic distances. We found barcode gaps for 72%-89% of taxa depending on the marker or markers used. With the exception of the markers rbcL and ITS, we found that evolutionary relatedness was an important predictor of barcode-gap presence or absence for all of the markers in combination and for matK, trnL-F, and trnH-psbA individually. This plant DNA barcode library and community phylogeny will be a valuable resource for future investigations.