Abstract
Recently reported insect declines have raised both political and social concern. Although the declines have been attributed to land use and climate change, supporting evidence suffers from ...low taxonomic resolution, short time series, a focus on local scales, and the collinearity of the identified drivers. In this study, we conducted a systematic assessment of insect populations in southern Germany, which showed that differences in insect biomass and richness are highly context dependent. We found the largest difference in biomass between semi-natural and urban environments (−42%), whereas differences in total richness (−29%) and the richness of threatened species (−56%) were largest from semi-natural to agricultural environments. These results point to urbanization and agriculture as major drivers of decline. We also found that richness and biomass increase monotonously with increasing temperature, independent of habitat. The contrasting patterns of insect biomass and richness question the use of these indicators as mutual surrogates. Our study provides support for the implementation of more comprehensive measures aimed at habitat restoration in order to halt insect declines.
Beetles are the most diverse group of animals and are crucial for ecosystem functioning. In many countries, they are well established for environmental impact assessment, but even in the well‐studied ...Central European fauna, species identification can be very difficult. A comprehensive and taxonomically well‐curated DNA barcode library could remedy this deficit and could also link hundreds of years of traditional knowledge with next generation sequencing technology. However, such a beetle library is missing to date. This study provides the globally largest DNA barcode reference library for Coleoptera for 15 948 individuals belonging to 3514 well‐identified species (53% of the German fauna) with representatives from 97 of 103 families (94%). This study is the first comprehensive regional test of the efficiency of DNA barcoding for beetles with a focus on Germany. Sequences ≥500 bp were recovered from 63% of the specimens analysed (15 948 of 25 294) with short sequences from another 997 specimens. Whereas most specimens (92.2%) could be unambiguously assigned to a single known species by sequence diversity at CO1, 1089 specimens (6.8%) were assigned to more than one Barcode Index Number (BIN), creating 395 BINs which need further study to ascertain if they represent cryptic species, mitochondrial introgression, or simply regional variation in widespread species. We found 409 specimens (2.6%) that shared a BIN assignment with another species, most involving a pair of closely allied species as 43 BINs were involved. Most of these taxa were separated by barcodes although sequence divergences were low. Only 155 specimens (0.97%) show identical or overlapping clusters.
This study presents DNA barcode records for 4118 specimens representing 561 species of bees belonging to the six families of Apoidea (Andrenidae, Apidae, Colletidae, Halictidae, Megachilidae and ...Melittidae) found in Central Europe. These records provide fully compliant barcode sequences for 503 of the 571 bee species in the German fauna and partial sequences for 43 more. The barcode results are largely congruent with traditional taxonomy as only five closely allied pairs of species could not be discriminated by barcodes. As well, 90% of the species possessed sufficiently deep sequence divergence to be assigned to a different Barcode Index Number (BIN). In fact, 56 species (11%) were assigned to two or more BINs reflecting the high levels of intraspecific divergence among their component specimens. Fifty other species (9.7%) shared the same Barcode Index Number with one or more species, but most of these species belonged to a distinct barcode cluster within a particular BIN. The barcode data contributed to clarifying the status of nearly half the examined taxonomically problematic species of bees in the German fauna. Based on these results, the role of DNA barcoding as a tool for current and future taxonomic work is discussed.
The German Barcoding initiatives BFB and GBOL have generated a reference library of more than 16,000 metazoan species, which is now ready for applications concerning next generation molecular ...biodiversity assessments. To streamline the barcoding process, we have developed a meta-barcoding pipeline: We pre-sorted a single malaise trap sample (obtained during one week in August 2014, southern Germany) into 12 arthropod orders and extracted DNA from pooled individuals of each order separately, in order to facilitate DNA extraction and avoid time consuming single specimen selection. Aliquots of each ordinal-level DNA extract were combined to roughly simulate a DNA extract from a non-sorted malaise sample. Each DNA extract was amplified using four primer sets targeting the CO1-5' fragment. The resulting PCR products (150-400bp) were sequenced separately on an Illumina Mi-SEQ platform, resulting in 1.5 million sequences and 5,500 clusters (coverage ≥10; CD-HIT-EST, 98%). Using a total of 120,000 DNA barcodes of identified, Central European Hymenoptera, Coleoptera, Diptera, and Lepidoptera downloaded from BOLD we established a reference sequence database for a local CUSTOM BLAST. This allowed us to identify 529 Barcode Index Numbers (BINs) from our sequence clusters derived from pooled Malaise trap samples. We introduce a scoring matrix based on the sequence match percentages of each amplicon in order to gain plausibility for each detected BIN, leading to 390 high score BINs in the sorted samples; whereas 268 of these high score BINs (69%) could be identified in the combined sample. The results indicate that a time consuming presorting process will yield approximately 30% more high score BINs compared to the non-sorted sample in our case. These promising results indicate that a fast, efficient and reliable analysis of next generation data from malaise trap samples can be achieved using this pipeline.
One key target of agri-environmental schemes (AES) is maintaining biodiversity. However, if and how insect biomass and diversity benefit from grassland AES is not well known. Using Malaise trap ...samples from 72 grassland sites in four grassland dominated regions in Southern Germany, we investigated the effect of 1) local grassland AES (three levels: control, farm-wide, site-specific), and 2) an index of land use extensification (LEI, calculated for each site) on insect biomass and species richness of all insects, of five insect orders including all species (Coleoptera, Diptera, Hemiptera, Hymenoptera, Lepidoptera), and two pollinator groups (Syrphidae, Apiformes). Moreover, the effect of grassland AES in the surrounding landscape was quantified. Insect samples were subjected to metabarcoding to create DNA barcode sequences which were then compared with reference libraries (BOLD; NCBI) and additionally classified using the Ribosomal Database Project (RDP) classifier. Resulting taxonomies were combined into a consensus taxonomy table and the potential minimum number of species per sample was estimated. Statistical analyses were carried out using Generalized linear mixed models and Bayesian inference. Our results indicated that species richness of many insect groups benefited from AES. As predicted by the regression model, presence of site-specific grassland AES increased the species richness of all insects (∼8%), Hymenoptera (∼27%), Lepidoptera (∼35%) and Syrphidae (∼26%). While Apiformes benefited most, we found an average increase of species richness in the presence of farm-wide AES by ∼82% and even more with site-specific AES ∼120%. We found that an increasing LEI, and thus more extensive management, positively affected insect diversity in grassland habitats. No effect of local AES and LEI on insect biomass was found. Moreover, the amount of grassland AES in the landscape was less important for insect diversity. We conclude that many grassland insects (especially pollinators) benefit from extensive management. However, we showed that the studied AES variants comprise variable management intensities hampering general AES recommendations for promoting insects. AES programs should, therefore, specifically reward extensification measures that were shown to benefit insect diversity. The integration of the land use extensification index (LEI) as a tool in the AES framework seems a promising approach to preserve and promote of insect diversity.
•Site-specific agri-environmental schemes enhance insect diversity in grasslands.•Extensification drives insect diversity, but not insect biomass.•Extensification effects differ among insect taxa with Apiformes benefiting most.•Amount of grassland AES in the landscape was less important for insect diversity.•Land use extensification index is proposed as a tool for targeted future AES.
As part of the German Barcode of Life campaign, over 3500 arachnid specimens have been collected and analyzed: ca. 3300 Araneae and 200 Opiliones, belonging to almost 600 species (median: 4 ...individuals/species). This covers about 60% of the spider fauna and more than 70% of the harvestmen fauna recorded for Germany. The overwhelming majority of species could be readily identified through DNA barcoding: median distances between closest species lay around 9% in spiders and 13% in harvestmen, while in 95% of the cases, intraspecific distances were below 2.5% and 8% respectively, with intraspecific medians at 0.3% and 0.2%. However, almost 20 spider species, most notably in the family Lycosidae, could not be separated through DNA barcoding (although many of them present discrete morphological differences). Conspicuously high interspecific distances were found in even more cases, hinting at cryptic species in some instances. A new program is presented: DiStats calculates the statistics needed to meet DNA barcode release criteria. Furthermore, new generic COI primers useful for a wide range of taxa (also other than arachnids) are introduced.
Prey spectra (the number and composition of captured arthropods) represent a crucial aspect of carnivorous plant ecology, yet remain poorly studied. Traditional morphology-based approaches for prey ...identification are time-intensive, require specialists with considerable knowledge of arthropod taxonomy, and are hampered by high numbers of unidentifiable (i.e., heavily digested) prey items. We examined prey spectra of three species of closely-related annual Drosera (Droseraceae, sundews) from tropical northern Australia using a novel DNA metabarcoding approach with in-situ macro photography as a plausibility control and to facilitate prey quantity estimations. This new method facilitated accurate analyses of carnivorous plant prey spectra (even of heavily digested prey lacking characteristic morphological features) at a taxonomic resolution and level of completeness far exceeding morphology-based methods and approaching the 100% mark at arthropod order level. Although the three studied species exhibited significant differences in detected prey spectra, little prey specialisation was observed and habitat or plant population density variations were likely the main drivers of prey spectra dissimilarity.
The ‘Symphyta’ is a paraphyletic assemblage at the base of the order Hymenoptera, comprising 14 families and about 8750 species. All have phytophagous larvae, except for the Orussidae, which are ...parasitoids. This study presents and evaluates the results of DNA barcoding of approximately 5360 specimens of ‘Symphyta’, mainly adults, and 4362 sequences covering 1037 species were deemed of suitable quality for inclusion in the analysis. All extant families are represented, except for the Anaxyelidae. The majority of species and specimens are from Europe, but approximately 38% of the species and 13% of the specimens are of non‐European origin. The utility of barcoding for species identification and taxonomy of ‘Symphyta’ is discussed on the basis of examples from each of the included families. A significant level of cryptic species diversity was apparent in many groups. Other attractive applications include the identification of immature stages without the need to rear them, community analyses based on metabarcoding of bulk samples and association of the sexes of adults.
Mayflies, stoneflies and caddisflies (Ephemeroptera, Plecoptera and Trichoptera) are prominent representatives of aquatic macroinvertebrates, commonly used as indicator organisms for water quality ...and ecosystem assessments. However, unambiguous morphological identification of EPT species, especially their immature life stages, is a challenging, yet fundamental task. A comprehensive DNA barcode library based upon taxonomically well-curated specimens is needed to overcome the problematic identification. Once available, this library will support the implementation of fast, cost-efficient and reliable DNA-based identifications and assessments of ecological status. This study represents a major step towards a DNA barcode reference library as it covers for two-thirds of Germany's EPT species including 2,613 individuals belonging to 363 identified species. As such, it provides coverage for 38 of 44 families (86%) and practically all major bioindicator species. DNA barcode compliant sequences (≥500 bp) were recovered from 98.74% of the analysed specimens. Whereas most species (325, i.e., 89.53%) were unambiguously assigned to a single Barcode Index Number (BIN) by its COI sequence, 38 species (18 Ephemeroptera, nine Plecoptera and 11 Trichoptera) were assigned to a total of 89 BINs. Most of these additional BINs formed nearest neighbour clusters, reflecting the discrimination of geographical subclades of a currently recognized species. BIN sharing was uncommon, involving only two species pairs of Ephemeroptera. Interestingly, both maximum pairwise and nearest neighbour distances were substantially higher for Ephemeroptera compared to Plecoptera and Trichoptera, possibly indicating older speciation events, stronger positive selection or faster rate of molecular evolution.
The Lut Desert, southeastern Iran, regularly has the largest contiguous area of surface temperatures above 65°C compared to anywhere else on Earth. Despite these harsh conditions, some specialized ...species thrive in this extreme habitat. Misonne's spider gecko, Rhinogecko misonnei de Witte, 1973 (Reptilia, Squamata, Gekkonidae) is an endemic species that is native to the central part of the Lut Desert. This research aimed to explore the dietary gut content and ecology of Misonne's spider gecko populations inhabiting the Lut Desert. A DNA metabarcoding approach was applied to obtain data from the stomach content of six collected specimens. A total of 147,451 amplicon reads representing a total of 94 OTU (=Operational Taxonomic Units) for the CO1‐5P barcode region were sequenced. Of 94 OTUs, 50 (53.2%) identically matched ≥97% with BOLD and/or NCBI database and were identified at the genus level. Blasting in both databases revealed the presence of three phyla including Chordata (55.23%), Arthropoda (44.72%), and Annelida (0.05). 99.54% of Chordata DNA belonged to Rhinogecko misonnei mainly coming from stomach tissue of the examined specimens. Within the arthropods, 87.64% of the OTUs matched Insecta and 12.08% Arachnida. Our findings indicate that Rhinogecko misonnei is a generalist gecko that feeds on a variety of food items. About 81% of Rhinogecko food items in the central Lut are migratory insects that come from adjacent areas of the Lut District, although the arachnid food items originate from the central Lut. This study emphasizes the importance of the insect fauna of adjacent areas of the Lut Desert for the stability of the Desert food web.
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