A metabarcoding assessment of the diet of the insectivorous bats of Madeira Island, Macaronesia.

Understanding the trophic structure of species assemblages is crucial in order to comprehend how syntropic species coexist in space and time. Bats are the second most taxonomically diverse group of mammals and display a wide range of dietary strategies. Due to their ability to disperse over water, ca. 60% of all extant bat species occur on islands and for the most part their interspecific ecological interactions are poorly known. Using DNA metabarcoding, this study offers the first insights into the diet of Macaronesian bats by providing a holistic overview of prey consumed by all 3 bat species found on Madeira Island (Pipistrellus maderensis, Nyctalus leisleri verrucosus, and Plecotus austriacus) and investigating both interspecific (between P. maderensis and N. l. verrucosus) and intraspecific (between female and male N. l. verrucosus) dietary differences. We identified a total of 110 species of arthropod prey in the diet of the 3 bat species, including multiple agriculture and forestry pest species, a human disease-relevant species, and numerous taxa not previously recorded on the island. Lepidoptera was the primary prey order for all 3 bat species. The diet composition of P. maderensis and N. l. verrucosus differed significantly, with P. maderensis consuming more Diptera and multiple prey taxa not found in the diet of N. l. verucosus. Moreover, male N. l. verrucosus exhibited a broader niche breadth than females. This study is among the first to use DNA metabarcoding to evaluate the diet of insular bats and thus greatly advances knowledge regarding the trophic ecology and pest suppression services of these poorly-known mammals.

The InBIO Barcoding Initiative Database: DNA barcodes of Orthoptera from Portugal.

Background: The InBIO Barcoding Initiative (IBI) Orthoptera dataset contains records of 420 specimens covering all the eleven Orthoptera families occurring in Portugal. Specimens were collected in continental Portugal from 2005 to 2021 and were morphologically identified to species level by taxonomists. A total of 119 species were identified corresponding to about 77% of all the orthopteran species known from continental Portugal. New information: DNA barcodes of 54 taxa were made public for the first time at the Barcode of Life Data System (BOLD). Furthermore, the submitted sequences were found to cluster in 129 BINs (Barcode Index Numbers), 35 of which were new additions to the Barcode of Life Data System (BOLD). All specimens have their DNA barcodes publicly accessible through BOLD online database. Stenobothruslineatus is recorded for the first time for continental Portugal. This dataset greatly increases the knowledge on the DNA barcodes and distribution of Orthoptera from Portugal. All DNA extractions and most specimens are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources.

Dietary metabarcoding reveals the simplification of bird-pest interaction networks across a gradient of agricultural cover.

Agriculture is vital for supporting human populations, but its intensification often leads to landscape homogenization and a decline in non-provisioning ecosystem services. Ecological intensification and multifunctional landscapes are suggested as nature-based alternatives to intensive agriculture, using ecological processes like natural pest regulation to maximize food production. Birds are recognized for their role in increasing crop yields by consuming invertebrate pests in several agroecosystems. However, the understanding of how bird species, their traits and agricultural land cover influence the structure of bird-pest interactions remains limited. We sampled bird-pest interactions monthly for 1 year, at four sites within a multifunctional landscape, following a gradient of increasing agricultural land cover. We analysed 2583 droppings of 55 bird species with DNA metabarcoding and detected 225 pest species in 1139 samples of 42 bird species. As expected, bird-pest interactions were highly variable across bird species. Dietary pest richness was lower in the fully agricultural site, while predation frequency remained consistent across the agricultural land cover gradient. Network analysis revealed a reduction in the complexity of bird-pest interactions as agricultural coverage increased. Bird species abundance affected the bird's contribution to the network structure more than any of the bird traits analysed (weight, phenology, invertebrate frequency in diet and foraging strata), with more common birds being more important to network structure. Overall, our results show that increasing agricultural land cover increases the homogenization of bird-pest interactions. This shows the importance of maintaining natural patches within agricultural landscapes for biodiversity conservation and enhanced biocontrol.

A agricultura é essencial para suportar a população humana, mas a sua intensificação geralmente leva à homogeneização da paisagem e à redução dos serviços do ecossistema que não sejam de provisão. A intensificação ecológica e paisagens multifuncionais são sugeridas como alternativas naturais à agricultura intensiva, utilizando processos ecológicos como a regulação natural de pragas para maximizar a produção de alimentos. As aves são conhecidas pelo seu papel no aumento da produtividade das culturas por consumirem pragas em diversos agroecossistemas. Contudo, o conhecimento de como as espécies de aves, as suas características e a cobertura agrícola influenciam as interações entre aves e pragas são limitados. Nós amostrámos estas interações mensalmente durante um ano, em quatro locais, numa paisagem multifuncional, ao longo um gradiente de aumento da cobertura agrícola. Analisamos 2583 dejetos de 55 espécies de aves com DNA metabarcoding e detetamos 225 espécies praga em 1139 amostras de 42 espécies de aves. Como esperado, as interações entre aves e pragas foram muito distintas entre as várias espécies de aves. A riqueza de pragas na dieta foi menor no local completamente dominado por área agrícola, enquanto a frequência de predação de pragas foi constante ao longo do gradiente de cobertura agrícola. A análise de redes demonstrou uma redução na complexidade das interações entre aves e pragas à medida que a cobertura agrícola aumenta. A abundância das espécies de aves influenciou mais a contribuição das aves para a estrutura da rede do que qualquer uma das características analisadas (peso, fenologia, frequência de invertebrados na dieta e estrato de alimentação), sendo as aves mais comuns as mais importantes na estrutura da rede. De forma geral, os nossos resultados indicam que o aumento da cobertura agrícola aumenta a homogeneização das interações entre aves e pragas. Isto demonstra a importância de preservar áreas naturais em paisagem agrícolas para a conservação de biodiversidade e melhor controlo biológico.

The InBIO Barcoding Initiative Database: DNA barcodes of Iberian Bees.

Background: Bees are important actors in terrestrial ecosystems and are recognised for their prominent role as pollinators. In the Iberian Peninsula, approximately 1,100 bee species are known, with nearly 100 of these species being endemic to the Peninsula. A reference collection of DNA barcodes, based on morphologically identified bee specimens, representing 514 Iberian species, was constructed. The "InBIO Barcoding Initiative Database: DNA Barcodes of Iberian bees" dataset contains records of 1,059 sequenced specimens. The species of this dataset correspond to about 47% of Iberian bee species diversity and 21% of endemic species diversity. For peninsular Portugal only, the corresponding coverage is 71% and 50%. Specimens were collected between 2014 and 2022 and are deposited in the research collection of Thomas Wood (Naturalis Biodiversity Center, The Netherlands), in the FLOWer Lab collection at the University of Coimbra (Portugal), in the Andreia Penado collection at the Natural History and Science Museum of the University of Porto (MHNC-UP) (Portugal) and in the InBIO Barcoding Initiative (IBI) reference collection (Vairão, Portugal). New information: Of the 514 species sequenced, 75 species from five different families are new additions to the Barcode of Life Data System (BOLD) and 112 new BINs were added. Whilst the majority of species were assigned to a single BIN (94.9%), 27 nominal species were assigned to multiple BINs. Although the placement into multiple BINs may simply reflect genetic diversity and variation, it likely also represents currently unrecognised species-level diversity across diverse taxa, such as Amegillaalbigena Lepeletier, 1841, Andrenarussula Lepeletier, 1841, Lasioglossumleucozonium (Schrank, 1781), Nomadafemoralis Morawitz, 1869 and Sphecodesalternatus Smith, 1853. Further species pairs of Colletes, Hylaeus and Nomada were placed into the same BINs, emphasising the need for integrative taxonomy within Iberia and across the Mediterranean Basin more broadly. These data substantially contribute to our understanding of bee genetic diversity and DNA barcodes in Iberia and provide an important baseline for ongoing taxonomic revisions in the West Palaearctic biogeographical region.

Diet of the critically endangered blackchin guitarfish Glaucostegus cemiculus revealed using DNA metabarcoding.

We present the first assessment of the diet of the blackchin guitarfish Glaucostegus cemiculus (Geoffroy Saint-Hilaire, 1817) for West Africa using DNA metabarcoding on stomach contents of individuals captured in the Bijagós Archipelago, Guinea-Bissau. The diet was dominated by crustaceans, particularly caramote prawn Penaeus kerathurus (frequency of occurrence [FO] = 74%, numerical frequency [NF] = 54%) and fiddler crab Afruca tangeri (FO = 74%, NF = 12%). Bony fishes were present in 30% of the stomachs. We highlight the importance of conservation action for intertidal habitats and their associated benthic invertebrates for the survival of the critically endangered blackchin guitarfish.

DNA metabarcoding, diversity partitioning and null models reveal mechanisms of seasonal trophic specialization in a Mediterranean warbler.

Optimal Foraging Theory (OFT) predicts that a population's trophic niche expansion should occur in periods of food scarcity as individuals begin to opportunistically exploit sub-optimal food items. However, the Niche Variation Hypothesis (NVH) posits that niche widening may result from increased among-individual differentiation due to food partitioning to avoid competition. We tested these hypotheses through a DNA metabarcoding study of the Sardinian Warbler (Curruca melanocephala) diet over a year. We used null models and the decomposition of beta diversity on among-individual dietary differentiation to infer the mechanisms driving the population's niche variation. Warblers fed frequently on berries, with a peak in late summer and, to a lesser extent, in autumn. Their diet also included a wide range of arthropods, with their prevalence varying among seasons. Consistent with OFT, the population's niche width was narrower in spring/summer when the population was strongly specialized in berries. In winter, the population's niche expanded, possibly reflecting seasonal declines in food abundance. As predicted by NVH, among-individual differentiation tended to be higher in winter, but this was mainly due to increased differences in dietary richness rather than to the partitioning of resources. Overall, our results suggest that within-individual niche does not increase in lean periods, and instead, individuals adopt either a more opportunistic or more specialized foraging strategy. Increased competition in periods of scarcity may help explain such patterns, but instead of showing increased food partitioning as expected from NVH, it may reflect OFT mechanisms on individuals with differential competitive ability to access better food resources.

DNA Barcode library of the endemic-rich avifauna of the oceanic islands of the Gulf of Guinea.

Background: The BioSTP: DNA Barcoding of endemic birds from oceanic islands of the Gulf of Guinea dataset contains records of 155 bird specimens belonging to 56 species in 23 families, representing over 80% of the diversity of the breeding landbird community. All specimens were collected on Príncipe, São Tomé and Annobón Islands between 2002 and 2021 and morphologically identified to species or subspecies level by qualified ornithologists. The dataset includes all endemic species and 3/4 of the extant endemic subspecies of the islands. This dataset is the second release by BioSTP and it greatly increases the knowledge on the DNA barcodes of Gulf of Guinea birds. All DNA extractions are deposited at Associação BIOPOLIS - CIBIO, Research Center in Biodiversity and Genetic Resources. New information: The dataset includes DNA barcodes for all 29 endemic bird species and for 11 of the 15 extant endemic bird subspecies from the oceanic islands of the Gulf of Guinea. This is the first major DNA barcode set of African birds. The three endemic subspecies of Crithagrarufobrunnea, an island endemic with three allopatric populations within the Archipelago, are also represented. Additionally, we obtained DNA barcodes for 16 of the 21 non-endemic landbirds and for one vagrant (Sylviacommunis). In total, forty-one taxa were new additions to the Barcode of Life Data System (BOLD), with another 11 corresponding to under-represented taxa in BOLD. Furthermore, the submitted sequences were found to cluster in 55 Barcode Index Numbers (BINs), 37 of which were new to BOLD. All specimens have their DNA barcodes publicly accessible through BOLD online database and GenBank.

A species-level trait dataset of bats in Europe and beyond.

Knowledge of species' functional traits is essential for understanding biodiversity patterns, predicting the impacts of global environmental changes, and assessing the efficiency of conservation measures. Bats are major components of mammalian diversity and occupy a variety of ecological niches and geographic distributions. However, an extensive compilation of their functional traits and ecological attributes is still missing. Here we present EuroBaTrait 1.0, the most comprehensive and up-to-date trait dataset covering 47 European bat species. The dataset includes data on 118 traits including genetic composition, physiology, morphology, acoustic signature, climatic associations, foraging habitat, roost type, diet, spatial behaviour, life history, pathogens, phenology, and distribution. We compiled the bat trait data obtained from three main sources: (i) a systematic literature and dataset search, (ii) unpublished data from European bat experts, and (iii) observations from large-scale monitoring programs. EuroBaTrait is designed to provide an important data source for comparative and trait-based analyses at the species or community level. The dataset also exposes knowledge gaps in species, geographic and trait coverage, highlighting priorities for future data collection.

DNA metabarcoding to assess prey overlap between tuna and seabirds in the Eastern tropical Atlantic: Implications for an ecosystem-based management.

Overfishing has been drastically changing food webs in marine ecosystems, and it is pivotal to quantify these changes at the ecosystem level. This is especially important for ecosystems with a high diversity of top predators such as the Eastern Atlantic marine region. In this work we used high-throughput sequencing methods to describe the diet of the two most abundant tuna species, the Skipjack tuna (Katsuwonus pelamis) and the Yellowfin tuna (Thunnus albacares), highly targeted by fisheries off west Africa. We also explored prey diversity overlap between these tuna species and the seabird species breeding in Cabo Verde that are most likely to share prey preferences and suffer from bycatch, the Brown booby (Sula leucogaster) and Cape Verde shearwater (Calonectris edwardsii). Overall, the diet of both tuna species was more diverse than that of seabirds. Skipjack tuna diet was dominated by prey from lower trophic levels, such as krill, anchovies, and siphonophores, while the Yellowfin tuna diet was mainly based on epipelagic fish such as flying and halfbeak fishes. Some of the most abundant prey families detected in the Yellowfin tuna diet were shared with both seabird species, resulting in a high prey diversity overlap between this tuna species and seabirds These results have implications for the management of tuna fisheries in the Eastern Tropical Atlantic, because a large decrease of both tuna species might have cascading effects on both primary and secondary consumer levels, and the decrease of these underwater predators may have implications on the viability of tropical seabird populations.

The InBIO Barcoding Initiative Database: contribution to the knowledge on DNA barcodes of cuckoo wasps, with the description of new species from the Iberian Peninsula (Hymenoptera, Chrysididae).

Background: DNA barcoding technologies have provided a powerful tool for the fields of ecology and systematics. Here, we present a part of the InBIO Barcoding Initiative Database: contribution to the knowledge on DNA barcodes of cuckoo wasps (Hymenoptera, Chrysididae) dataset representing 144 specimens and 103 species, covering approximately 44% of the Iberian and 21% of the European fauna. The InBIO Barcoding Initiative (IBI - DNA Barcoding Portuguese terrestrial invertebrate biodiversity) aims to fill the barcoding gap for the terrestrial invertebrate taxa. All DNA extractions are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources and specimens are deposited in the University of Mons collection (Belgium) and in the Natur-Museum in Lucerne (Switzerland). New information: This dataset increases the knowledge on the DNA barcodes and distribution of 102 species of cuckoo wasps. A total of 52 species, from 11 different genera, were new additions to the Barcode of Life Data System (BOLD), with DNA barcodes for another 44 species added from under-represented taxa in BOLD. All specimens have their DNA barcodes publicly accessible through the BOLD online database. Nine cuckoo wasp species are newly recorded for Portugal. Additionally, two new species for science are described: Chrysiscrossi Rosa, sp. nov. from southern Portugal and Hedychridiumcalcarium Rosa, sp. nov. from eastern Spain. Several taxonomic changes are proposed and Hedychrumrutilans Dahlbom, 1845 is found to consist of two different taxa that can be found in sympatry, Hedychrumrutilans s. str. and Hedychrumviridaureum Tournier, 1877 stat. nov. Stilbumwestermanni Dahlbom, 1845 stat. nov. is confirmed as distinct from Stilbumcalens (Fabricius, 1781), with the latter species not confirmed as present in Iberia; barcoded Stilbum material from Australia is distinct and represents Stilbumamethystium (Fabricius, 1775) sp. resurr.; Portuguese material identified as Hedychridiumchloropygum Buysson, 1888 actually belongs to Hedychridiumcaputaureum Trautmann & Trautmann, 1919, the first confirmed record of this species from Iberia. Philoctetesparvulus (Dahlbom, 1845) is confirmed to be a synonym of Philoctetespunctulatus (Dahlbom, 1845). Chrysislusitanica Bischoff, 1910 is confirmed as a valid species. Chrysishebraeica Linsenmaier, 1959 stat. nov. is raised to species status.

Crowding after sudden habitat loss affects demography and social structure in a bat population.

The sudden loss of habitats due to natural or anthropogenic disturbances causes displacement of mobile animals from affected areas to refuge habitats, where large but often transitory concentrations of individuals may occur. While these local density increases have been previously described, the hypothesis that crowding disrupts demographic processes remains largely untested. Here we used the sudden flooding of a river valley by a hydroelectric reservoir as a quasi-experiment to investigate the consequences of crowding on demography, fecundity and social structure in the European free-tailed bat Tadarida teniotis. We monitored bat populations at roosts near and far from the flooded area, before (2013-2014), during (2015) and after (2016) habitat flooding. We assessed population demographic parameters using Capture-Mark-Recapture (CMR) models (3,821 PIT-tagged individuals), and used genetic relatedness among individuals (1,407 individuals genotyped for 14 microsatellite markers) to infer changes in social structure. Habitat loss through flooding was associated with significant but transitory increases in the number of bats using nearby roosts. This may be related to the higher probability of individuals arriving at those roosts during flooding, together with increases in individual local residency through time, particularly among males. Individual apparent survival was highest during flooding and lowest in the following year, while the probability of leaving a roost safe from flooding was higher near the impact area than farther away. Crowding did not negatively affect fecundity, but the arrival of new individuals led to changes in social structure as revealed by lower genetic relatedness between individuals after disturbance at roosts near the flooding area, but not in those farther afield. Our study documents a clear example of crowding effects, suggesting that bats losing roosts due to a hydroelectric reservoir moved to alternative roosts, where local increases in population size and the arrival of new individuals reduced genetic relatedness and apparent survival, but not fecundity. These results support the hypothesis that crowding after habitat loss can disrupt population processes, even though effects may be subtle and short-lived. Also, they point out the need to duly consider crowding effects when assessing and mitigating anthropogenic impacts on animal populations.

The InBIO Barcoding Initiative Database: DNA barcodes of Portuguese Diptera 02 - Limoniidae, Pediciidae and Tipulidae.

BACKGROUND: The InBIO Barcoding Initiative (IBI) Diptera 02 dataset contains records of 412 crane fly specimens belonging to the Diptera families: Limoniidae, Pediciidae and Tipulidae. This dataset is the second release by IBI on Diptera and it greatly increases the knowledge on the DNA barcodes and distribution of crane flies from Portugal. All specimens were collected in Portugal, including six specimens from the Azores and Madeira archipelagos. Sampling took place from 2003 to 2019. Specimens have been morphologically identified to species level by taxonomists and belong to 83 species in total. The species, represented in this dataset, correspond to about 55% of all the crane fly species known from Portugal and 22% of crane fly species known from the Iberian Peninsula. All DNA extractions and most specimens are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources. NEW INFORMATION: Fifty-three species were new additions to the Barcode of Life Data System (BOLD), with another 18 species' barcodes added from under-represented species in BOLD. Furthermore, the submitted sequences were found to cluster in 88 BINs, 54 of which were new to BOLD. All specimens have their DNA barcodes publicly accessible through BOLD online database and its collection data can be accessed through the Global Biodiversity Information Facility (GBIF). One species, Gonomyiatenella (Limoniidae), is recorded for the first time from Portugal, raising the number of crane flies recorded in the country to 145 species.

Combining DNA metabarcoding and ecological networks to inform conservation biocontrol by small vertebrate predators.

In multifunctional landscapes, diverse communities of flying vertebrate predators provide vital services of insect pest control. In such landscapes, conservation biocontrol should benefit service-providing species to enhance the flow, stability and resilience of pest control services supporting the production of food and fiber. However, this would require identifying key service providers, which may be challenging when multiple predators interact with multiple pests. Here we provide a framework to identify the functional role of individual species to pest control in multifunctional landscapes. First, we used DNA metabarcoding to provide detailed data on pest species predation by diverse predator communities. Then, these data were fed into an extensive network analysis, in which information relevant for conservation biocontrol is gained from parameters describing network structure (e.g., modularity) and species roles in such network (e.g., centrality, specialization). We applied our framework to a Mediterranean landscape, where 19 bat species were found to feed on 132 insect pest species. Metabarcoding data revealed potentially important bats that consumed insect pest species in high frequency and/or diversity. Network analysis showed a modular structure, indicating sets of bat species that are required to regulate specific sets of insect pests. A few generalist bats had particularly important roles, either at network or module levels. Extinction simulations highlighted six bats, including species of conservation concern, which were sufficient to ensure that over three-quarters of the pest species had at least one bat predator. Combining DNA metabarcoding and ecological network analysis provides a valuable framework to identify individual species within diverse predator communities that might have a disproportionate contribution to pest control services in multifunctional landscapes. These species can be regarded as candidate targets for conservation biocontrol, although additional information is needed to evaluate their actual effectiveness in pest regulation.

Low MSP-1 haplotype diversity in the West Palearctic population of the avian malaria parasite Plasmodium relictum.

BACKGROUND: Although avian Plasmodium species are widespread and common across the globe, limited data exist on how genetically variable their populations are. Here, the hypothesis that the avian blood parasite Plasmodium relictum exhibits very low genetic diversity in its Western Palearctic transmission area (from Morocco to Sweden in the north and Transcaucasia in the east) was tested. METHODS: The genetic diversity of Plasmodium relictum was investigated by sequencing a portion (block 14) of the fast-evolving merozoite surface protein 1 (MSP1) gene in 75 different P. relictum infections from 36 host species. Furthermore, the full-length MSP1 sequences representing the common block 14 allele was sequenced in order to investigate if additional variation could be found outside block 14. RESULTS: The majority (72 of 75) of the sequenced infections shared the same MSP1 allele. This common allele has previously been found to be the dominant allele transmitted in Europe. CONCLUSION: The results corroborate earlier findings derived from a limited dataset that the globally transmitted malaria parasite P. relictum exhibits very low genetic diversity in its Western Palearctic transmission area. This is likely the result of a recent introduction event or a selective sweep.

High-resolution multi-marker DNA metabarcoding reveals sexual dietary differentiation in a bird with minor dimorphism.

Although sexual dietary differentiation is well known in birds, it is usually linked with significant morphological dimorphism between males and females, with lower differentiation reported in sexually monomorphic or only slightly dimorphic species. However, this may be an artifact of poor taxonomic resolution achieved in most conventional dietary studies, which may be unable to detect subtle intraspecific differentiation in prey consumption. Here, we show the power of multi-marker metabarcoding to address these issues, focusing on a slightly dimorphic generalist passerine, the black wheatear Oenanthe leucura. Using markers from four genomic regions (18S, 16S, COI, and trnL), we analyzed fecal droppings collected from 93 adult black wheatears during the breeding season. We found that sexes were rather similar in bill and body features, though males had a slightly thicker bill and longer wings and tail than females. Diet was dominated in both sexes by a very wide range of arthropod species and a few fleshy fruits, but the overall diet diversity was higher for males than females, and there was a much higher frequency of occurrence of ants in female (58%) than male (29%) diets. We hypothesize that the observed sexual differentiation was likely related to females foraging closer to their offspring on abundant prey, while males consumed a wider variety of prey while foraging more widely. Overall, our results suggest that dietary sexual differentiation in birds may be more widespread than recognized at present and that multi-marker DNA metabarcoding is a particularly powerful tool to unveiling such differences.

Hidden in our pockets: building of a DNA barcode library unveils the first record of Myotis alcathoe for Portugal.

BACKGROUND: The advent and boom of DNA barcoding technologies have provided a powerful tool for the fields of ecology and systematics. Here, we present the InBIO Barcoding Initiative Database: Portuguese Bats (Chiroptera) dataset containing DNA sequences of 63 specimens representing the 25 bat species currently known for continental Portugal. For that, we sequenced tissues samples obtained in a vast array of projects spanning the last two decades. NEW INFORMATION: We added four new Barcoding Index Numbers (BINs) to existing Chiroptera barcodes on BOLD, two belonging to Myotis escalerai, one to Plecotus auritus and the other to Rhinolophus hipposideros. Surprisingly, one of the samples initially identified in the field as Myotis mystacinus turned out to be Myotis alcathoe, which represents the first record of this species for Portugal. The presence of Nyctalus noctula in Portugal was also genetically confirmed for the first time. This case study shows the power and value of DNA barcoding initiatives to unravel new data that may be hidden on biological collections.

Ypsolopha rhinolophi sp. nov. (Lepidoptera: Ypsolophidae), a new species from Portugal and France unveiled by bats.

A new species Ypsolopha rhinolophi Corley is described from northern Portugal and south-east France. It resembles Y. alpella (Denis Schiffermüller, 1775) and Y. lucella (Fabricius, 1775) but shows clear differences from both species in DNA barcode and in male and female genitalia. Male genitalia of Y. lucella are illustrated for the first time. The new species has been collected at light, reared from larvae on Quercus pyrenaica Willd. and recognised from DNA barcode fragments obtained from droppings of horseshoe bats.

Advancing the integration of multi-marker metabarcoding data in dietary analysis of trophic generalists.

The application of DNA metabarcoding to dietary analysis of trophic generalists requires using multiple markers in order to overcome problems of primer specificity and bias. However, limited attention has been given to the integration of information from multiple markers, particularly when they partly overlap in the taxa amplified, and vary in taxonomic resolution and biases. Here, we test the use of a mix of universal and specific markers, provide criteria to integrate multi-marker metabarcoding data and a python script to implement such criteria and produce a single list of taxa ingested per sample. We then compare the results of dietary analysis based on morphological methods, single markers, and the proposed combination of multiple markers. The study was based on the analysis of 115 faeces from a small passerine, the Black Wheatears (Oenanthe leucura). Morphological analysis detected far fewer plant taxa (12) than either a universal 18S marker (57) or the plant trnL marker (124). This may partly reflect the detection of secondary ingestion by molecular methods. Morphological identification also detected far fewer taxa (23) than when using 18S (91) or the arthropod markers IN16STK (244) and ZBJ (231), though each method missed or underestimated some prey items. Integration of multi-marker data provided far more detailed dietary information than any single marker and estimated higher frequencies of occurrence of all taxa. Overall, our results show the value of integrating data from multiple, taxonomically overlapping markers in an example dietary data set.

Intricate trophic links between threatened vertebrates confined to a small island in the Atlantic Ocean.

Trophic networks in small isolated islands are in a fragile balance, and their disturbance can easily contribute toward the extinction vortex of species. Here, we show, in a small Atlantic island (Raso) in the Cabo Verde Archipelago, using DNA metabarcoding, the extent of trophic dependence of the Endangered giant wall gecko Tarentola gigas on endemic populations of vertebrates, including one of the rarest bird species of the world, the Critically Endangered Raso lark Alauda razae. We found that the Raso lark (27%), Iago sparrow Passer iagoensis (12%), Bulwer's petrel Bulweria bulwerii (15%), and the Cabo Verde shearwater Calonectris edwardsii (10%) are the most frequent vertebrate signatures found in the feces of the giant wall gecko. This work provides the first integrative assessment of their trophic links, an important issue to be considered for the long-term conservation of these small and isolated island ecosystems.