Hidden diversity of Acoelomorpha revealed through metabarcoding

Animals with bilateral symmetry comprise the majority of the described species within Metazoa. However, the nature of the first bilaterian animal remains unknown. As most recent molecular phylogenies point to Xenacoelomorpha as the sister group to the rest of Bilateria, understanding their biology, ecology and diversity is key to reconstructing the nature of the last common bilaterian ancestor (Urbilateria). To date, sampling efforts have focused mainly on coastal areas, leaving potential gaps in our understanding of the full diversity of xenacoelomorphs. We therefore analysed 18S rDNA metabarcoding data from three marine projects covering benthic and pelagic habitats worldwide. Our results show that acoels have a greater richness in planktonic environments than previously described. Interestingly, we also identified a putative novel clade of acoels in the deep benthos that branches as sister group to the rest of Acoela, thus representing the earliest-branching acoel clade. Our data highlight deep-sea environments as an ideal habitat to sample acoels with key phylogenetic positions, which might be useful for reconstructing the early evolution of Bilateria.     

Winogradsky columns as a strategy to study typically rare microbial eukaryotes

Winogradsky columns have been widely used to study soil microbial communities, but the vast majority of those investigations have focused on the ecology and diversity of bacteria. In contrast, microbial eukaryotes (ME) have been regularly overlooked in studies based on experimental soil columns. Despite the recognized ecological relevance of ME in soil communities, investigations focused on ME diversity and the abundance of certain groups of interest are still scarce. In the present study, we used DNA metabarcoding (high-throughput sequencing of the V4 region of the 18S rRNA locus) to survey the ME diversity and abundance in an experimental Winogradsky soil column. Consistent with previous surveys in natural soils, our survey identified members of Cercozoa (Rhizaria; 31.2%), Apicomplexa and Ciliophora (Alveolata; 12.5%) as the predominant ME groups, but at particular depths we also detected the abundant presence of ME lineages that are typically rare in natural environments, such as members of the Vampyrellida (Rhizaria) and Breviatea (Amorphea). Our survey demonstrates that experimental soil columns are an efficient enrichment-culture approach that can enhance investigations about the diversity and ecology of ME in soils.     

DNA metabarcoding provides insights into seasonal diet variations in Chinese mole shrew (Anourosorex squamipes) with potential implications for evaluating crop impacts

Diet analysis of potential small mammals pest species is important for understanding feeding ecology and evaluating their impact on crops and stored foods. Chinese mole shrew (Anourosorex squamipes), distributed in Southwest China, has previously been reported as a farmland pest. Effective population management of this species requires a better understanding of its diet, which can be difficult to determine with high taxonomic resolution using conventional microhistological methods. In this study, we used two DNA metabarcoding assays to identify 38 animal species and 65 plant genera from shrew stomach contents, which suggest that A. squamipes is an omnivorous generalist. Earthworms are the most prevalent (>90%) and abundant (>80%) food items in the diverse diet of A. squamipes. Species of the Fabaceae (frequency of occurrence [FO]: 88%; such as peanuts) and Poaceae (FO: 71%; such as rice) families were the most common plant foods identified in the diet of A. squamipes. Additionally, we found a seasonal decrease in the diversity and abundance of invertebrate foods from spring and summer to winter. Chinese mole shrew has a diverse and flexible diet throughout the year to adapt to seasonal variations in food availability, contributing to its survival even when food resources are limited. This study provides a higher resolution identification of the diet of A. squamipes than has been previously described and is valuable for understanding shrew feeding ecology as well as evaluating possible species impacts on crops.     

Seasonal and spatial variability of zooplankton diversity in the Poyang Lake Basin using DNA metabarcoding

Freshwater ecosystems face multiple threats to their stability globally. Poyang Lake is the largest lake in China, but its habitat has been seriously degraded because of human activities and natural factors (e.g. climate change), resulting in a decline in freshwater biodiversity. Zooplankton are useful indicators of environmental stressors because they are sensitive to external perturbations. DNA metabarcoding is an approach that has gained significant traction by aiding ecosystem conservation and management. Here, the seasonal and spatial variability in the zooplankton diversity were analyzed in the Poyang Lake Basin using DNA metabarcoding. The results showed that the community structure of zooplankton exhibited significant seasonal and spatial variability using DNA metabarcoding, where the community structure was correlated with turbidity, water temperature, pH, total phosphorus, and chlorophyll‐a. These results indicated habitat variations affected by human activities and seasonal change could be the main driving factors for the variations of zooplankton community. This study also provides an important reference for the management of aquatic ecosystem health and conservation of aquatic biodiversity.     

Disease‐related population declines in bats demonstrate non‐exchangeability in generalist predators

The extent to which persisting species may fill the functional role of extirpated or declining species has profound implications for the structure of biological communities and ecosystem functioning. In North America, arthropodivorous bats are threatened on a continent‐wide scale by the spread of white‐nose syndrome (WNS), a disease caused by the fungus Pseudogymnoascus destructans. We tested whether bat species that display lower mortality from this disease can partially fill the functional role of other bat species experiencing population declines. Specifically, we performed high‐throughput amplicon sequencing of guano from two generalist predators: the little brown bat (Myotis lucifugus) and big brown bat (Eptesicus fuscus). We then compared changes in prey consumption before versus after population declines related to WNS. Dietary niches contracted for both species after large and abrupt declines in little brown bats and smaller declines in big brown bats, but interspecific dietary overlap did not change. Furthermore, the incidence and taxonomic richness of agricultural pest taxa detected in diet samples decreased following bat population declines. Our results suggest that persisting generalist predators do not necessarily expand their dietary niches following population declines in other predators, providing further evidence that the functional roles of different generalist predators are ecologically distinct.     

Field investigation‐ and dietary metabarcoding‐based screening of arthropods that prey on primary tea pests

Predatory natural enemies play key functional roles in biological control. Abundant predatory arthropod species have been recorded in tea plantation ecosystems. However, few studies have comprehensively evaluated the control effect of predatory arthropods on tea pests in the field. We performed a 1‐year field investigation and collected predatory arthropods and pests in the tea canopy. A total of 7931 predatory arthropod individuals were collected, and Coleosoma blandum (Araneae, Theridiidae) was the most abundant species in the studied tea plantation. The population dynamics between C. blandum and four main tea pest species (Aleurocanthus spiniferus, Empoasca onukii, Ectropis grisescens, and Scopula subpunctaria) were established using the individual number of predators and pests in each month. The results showed that C. blandum appeared to co‐occur in the tea canopy with A. spiniferus, Em. onukii, and Ec. grisescens in a longer period. The prey spectrum of C. blandum was further analyzed using DNA metabarcoding. Among prey species, A. spiniferus, Em. onukii, and Ec. grisescens were included, and the relative abundance and positive rates of target DNA fragments of A. spiniferus were greater than that of other two pests. Combined with the high dominance index of C. blandum, co‐occurrence between C. blandum and A. spiniferus in time and space and high positive rate and relative abundance of target DNA fragments of A. spiniferus, C. blandum was identified to prey on A. spiniferus, and C. blandum may be an important predator of A. spiniferus. Thus, C. blandum has potential as a biological control agent of A. spiniferus in an integrated pest management strategy.     

A DNA barcode library for 5,200 German flies and midges (Insecta: Diptera) and its implications for metabarcoding‐based biomonitoring

This study summarizes results of a DNA barcoding campaign on German Diptera, involving analysis of 45,040 specimens. The resultant DNA barcode library includes records for 2,453 named species comprising a total of 5,200 barcode index numbers (BINs), including 2,700 COI haplotype clusters without species‐level assignment, so called “dark taxa.” Overall, 88 out of 117 families (75%) recorded from Germany were covered, representing more than 50% of the 9,544 known species of German Diptera. Until now, most of these families, especially the most diverse, have been taxonomically inaccessible. By contrast, within a few years this study provided an intermediate taxonomic system for half of the German Dipteran fauna, which will provide a useful foundation for subsequent detailed, integrative taxonomic studies. Using DNA extracts derived from bulk collections made by Malaise traps, we further demonstrate that species delineation using BINs and operational taxonomic units (OTUs) constitutes an effective method for biodiversity studies using DNA metabarcoding. As the reference libraries continue to grow, and gaps in the species catalogue are filled, BIN lists assembled by metabarcoding will provide greater taxonomic resolution. The present study has three main goals: (a) to provide a DNA barcode library for 5,200 BINs of Diptera; (b) to demonstrate, based on the example of bulk extractions from a Malaise trap experiment, that DNA barcode clusters, labelled with globally unique identifiers (such as OTUs and/or BINs), provide a pragmatic, accurate solution to the “taxonomic impediment”; and (c) to demonstrate that interim names based on BINs and OTUs obtained through metabarcoding provide an effective method for studies on species‐rich groups that are usually neglected in biodiversity research projects because of their unresolved taxonomy.     

Molecular diet analysis finds an insectivorous desert bat community dominated by resource sharing despite diverse echolocation and foraging strategies

Interspecific differences in traits can alter the relative niche use of species within the same environment. Bats provide an excellent model to study niche use because they use a wide variety of behavioral, acoustic, and morphological traits that may lead to multi‐species, functional groups. Predatory bats have been classified by their foraging location (edge, clutter, open space), ability to use aerial hawking or substrate gleaning and echolocation call design and flexibility, all of which may dictate their prey use. For example, high frequency, broadband calls do not travel far but offer high object resolution while high intensity, low frequency calls travel further but provide lower resolution. Because these behaviors can be flexible, four behavioral categories have been proposed: (a) gleaning, (b) behaviorally flexible (gleaning and hawking), (c) clutter‐tolerant hawking, and (d) open space hawking. Many recent studies of diet in bats use molecular tools to identify prey but mainly focus on one or two species in isolation; few studies provide evidence for substantial differences in prey use despite the many behavioral, acoustic, and morphological differences. Here, we analyze the diet of 17 sympatric species in the Chihuahuan desert and test the hypothesis that peak echolocation frequency and behavioral categories are linked to differences in diet. We find no significant correlation between dietary richness and echolocation peak frequency though it spanned close to 100 kHz across species. Our data, however, suggest that bats which use both gleaning and hawking strategies have the broadest diets and are most differentiated from clutter‐tolerant aerial hawking species.     

Environmental factors and host genetic variation shape the fungal endophyte communities within needles of Scots pine (Pinus sylvestris)

To determine the role of environmental and host genetic factors in shaping fungal endophyte communities we used culturing and metabarcoding techniques to quantify fungal taxa within healthy Scots pine (Pinus sylvestris) needles in a 7-y old provenance-progeny trial replicated at three sites. Both methods revealed a community of ascomycete and basidiomycete taxa dominated by the needle pathogen Lophodermium seditiosum. Differences in fungal endophyte taxon composition and diversity indices were highly significant among trial sites. Within two sites, fungal endophyte communities varied significantly among provenances. Furthermore, the communities differed significantly among maternal families within provenances in 11/15 and 7/15 comparisons involving culture and metabarcoding data respectively. We conclude that both environmental and host genetic variation shape the fungal endophyte community of P. sylvestris needles.     

DNA metabarcoding of feces to infer summer diet of Pacific walruses

Environmental conditions in the Chukchi Sea are changing rapidly and may alter the abundance and distribution of marine species and their benthic prey. We used a metabarcoding approach to identify potentially important prey taxa from Pacific walrus (Odobenus rosmarus divergens) fecal samples (n = 87). Bivalvia was the most dominant class of prey (66% of all normalized counts) and occurred in 98% of the samples. Polychaeta and Gastropoda occurred in 70% and 62% of the samples, respectively. The remaining nine invertebrate classes comprised <21% of all normalized counts. The common occurrence of these three prey classes is consistent with examinations of walrus stomach contents. Despite these consistencies, biases in the metabarcoding approach to determine diet from feces have been highlighted in other studies and require further study, in addition to biases that may have arisen from our opportunistic sampling. However, this noninvasive approach provides accurate identification of prey taxa from degraded samples and could yield much-needed information on shifts in walrus diet in a rapidly changing Arctic.