DNA from soil mirrors plant taxonomic and growth form diversity

Ecosystems across the globe are threatened by climate change and human activities. New rapid survey approaches for monitoring biodiversity would greatly advance assessment and understanding of these threats. Taking advantage of next-generation DNA sequencing, we tested an approach we call metabarcoding: high-throughput and simultaneous taxa identification based on a very short (usually <100 base pairs) but informative DNA fragment. Short DNA fragments allow the use of degraded DNA from environmental samples. All analyses included amplification using plant-specific versatile primers, sequencing and estimation of taxonomic diversity. We tested in three steps whether degraded DNA from dead material in soil has the potential of efficiently assessing biodiversity in different biomes. First, soil DNA from eight boreal plant communities located in two different vegetation types (meadow and heath) was amplified. Plant diversity detected from boreal soil was highly consistent with plant taxonomic and growth form diversity estimated from conventional above-ground surveys. Second, we assessed DNA persistence using samples from formerly cultivated soils in temperate environments. We found that the number of crop DNA sequences retrieved strongly varied with years since last cultivation, and crop sequences were absent from nearby, uncultivated plots. Third, we assessed the universal applicability of DNA metabarcoding using soil samples from tropical environments: a large proportion of species and families from the study site were efficiently recovered. The results open unprecedented opportunities for large-scale DNA-based biodiversity studies across a range of taxonomic groups using standardized metabarcoding approaches.     

Blind assessment of vertebrate taxonomic diversity across spatial scales by clustering environmental DNA metabarcoding sequences

Human activities impact all ecosystems on Earth, which urges scientists to better understand biodiversity changes across temporal and spatial scales. Environmental DNA (eDNA) metabarcoding is a promising non-invasive method to assess species composition in a wide range of ecosystems. Yet, this method requires the completeness of a reference database, i.e. a list of DNA sequences attached to each species of the regional pool, which is rarely met. As an alternative, molecular operational taxonomic units (MOTUs) can be extracted as clusters of sequences. However, the extent to which the diversity of MOTUs can predict the diversity of species across spatial scales is unknown. Here, we used 196 samples along the Rhone river (France) for which the reference database is complete to assess whether a blind eDNA approach can reliably predict the ground-truth number of species at different spatial scales. Using the 12S rDNA teleo primer, we curated and clustered 60 million sequences into MOTUs using a new assembled bioinformatic pipeline. We show that stringent quality filters were necessary to remove artefact noise, notably MOTUs present in a single PCR replicate, which represented 55% of MOTUs (103). Post-clustering cleaning also removed 19 additional erroneous MOTUs and only discarded one truly present species. We then show that the diversity of retained fish MOTUs accurately predicted the local (α, r = 0.98) and regional (γ) ground-truth species diversity (67 MOTUs versus 63 species), but also the species dissimilarity between samples (β-diversity, r = 0.98). This work paves the way towards extending the use of eDNA metabarcoding in community ecology and biogeography despite major gaps in genetic reference databases.     

Marine water environmental DNA metabarcoding provides a comprehensive fish diversity assessment and reveals spatial patterns in a large oceanic area

Current methods for monitoring marine fish (including bony fishes and elasmobranchs) diversity mostly rely on trawling surveys, which are invasive, costly, and time‐consuming. Moreover, these methods are selective, targeting a subset of species at the time, and can be inaccessible to certain areas. Here, we used environmental DNA (eDNA), the DNA present in the water column as part of shed cells, tissues, or mucus, to provide comprehensive information about fish diversity in a large marine area. Further, eDNA results were compared to the fish diversity obtained in pelagic trawls. A total of 44 5 L‐water samples were collected onboard a wide‐scale oceanographic survey covering about 120,000 square kilometers in Northeast Atlantic Ocean. A short region of the 12S rRNA gene was amplified and sequenced through metabarcoding generating almost 3.5 million quality‐filtered reads. Trawl and eDNA samples resulted in the same most abundant species (European anchovy, European pilchard, Atlantic mackerel, and blue whiting), but eDNA metabarcoding resulted in more detected bony fish and elasmobranch species (116) than trawling (16). Although an overall correlation between fishes biomass and number of reads was observed, some species deviated from the common trend, which could be explained by inherent biases of each of the methods. Species distribution patterns inferred from eDNA metabarcoding data coincided with current ecological knowledge of the species, suggesting that eDNA has the potential to draw sound ecological conclusions that can contribute to fish surveillance programs. Our results support eDNA metabarcoding for broad‐scale marine fish diversity monitoring in the context of Directives such as the Common Fisheries Policy or the Marine Strategy Framework Directive.     

Environmental DNA metabarcoding primers for freshwater fish detection and quantification: In silico and in tanks

Environmental DNA (eDNA) techniques refer to utilizing the organisms’ DNA extracted from environment samples to genetically identify target species without capturing actual organisms. eDNA metabarcoding via high‐throughput sequencing can simultaneously detect multiple fish species from a single water sample, which is a powerful tool for the qualitative detection and quantitative estimates of multiple fish species. However, sequence counts obtained from eDNA metabarcoding may be influenced by many factors, of which primer bias is one of the foremost causes of methodological error. The performance of 18 primer pairs for COI, cytb, 12S rRNA, and 16S rRNA mitochondrial genes, which are all frequently used in fish eDNA metabarcoding, were evaluated in the current study. The ribosomal gene markers performed better than the protein‐coding gene markers during in silico screening, resulting in higher taxonomic coverage and appropriate barcode lengths. Four primer pairs—AcMDB07, MiFish‐U, Ve16S1, and Ve16S3—designed for various regions of the 12S and 16S rRNA genes were screened for tank metabarcoding in a case study targeting six freshwater fish species. The four primer pairs were able to accurately detect all six species in different tanks, while only MiFish‐U, Ve16S1, and Ve16S3 revealed a significant positive relationship between species biomass and read count for the pooled tank data. The positive relationship could not be found in all species within the tanks. Additionally, primer efficiency differed depending on the species while primer preferential species varied in different fish assemblages. This case study supports the potential for eDNA metabarcoding to assess species diversity in natural ecosystems and provides an alternative strategy to evaluate the performance of candidate primers before application of eDNA metabarcoding in natural ecosystems.     

Trophic resource partitioning drives fine‐scale coexistence in cryptic bat species

Understanding the processes that enable species coexistence has important implications for assessing how ecological systems will respond to global change. Morphology and functional similarity increase the potential for competition, and therefore, co‐occurring morphologically similar but genetically unique species are a good model system for testing coexistence mechanisms. We used DNA metabarcoding and high‐throughput sequencing to characterize for the first time the trophic ecology of two recently described cryptic bat species with parapatric ranges, Myotis escalerai and Myotis crypticus. We collected fecal samples from allopatric and sympatric regions and from syntopic and allotopic locations within the sympatric region to describe the diets both taxonomically and functionally and compare prey consumption with prey availability. The two bat species had highly similar diets characterized by high arthropod diversity, particularly Lepidoptera, Diptera and Araneae, and a high proportion of prey that is not volant at night, which points to extensive use of gleaning. Diet overlap at the prey item level was lower in syntopic populations, supporting trophic shift under fine‐scale co‐occurrence. Furthermore, the diet of M. escalerai had a marginally lower proportion of not nocturnally volant prey in syntopic populations, suggesting that the shift in diet may be driven by a change in foraging mode. Our findings suggest that fine‐scale coexistence mechanisms can have implications for maintaining broad‐scale diversity patterns. This study highlights the importance of including both allopatric and sympatric populations and choosing meaningful spatial scales for detecting ecological patterns. We conclude that a combination of high taxonomic resolution with a functional approach helps identify patterns of niche shift.     

Mantoniella beaufortii and Mantoniella baffinensis sp. nov. (Mamiellales,Mamiellophyceae), two new green algal species from the high arctic1

Members of the class Mamiellophyceae comprise species that can dominate picophytoplankton diversity in polar waters. Yet, polar species are often morphologically indistinguishable from temperate species, although clearly separated by molecular features. Here we examine four Mamiellophyceae strains from the Canadian Arctic. The 18S rRNA and Internal Transcribed Spacer 2 (ITS2) gene phylogeny place these strains within the family Mamiellaceae (Mamiellales, Mamiellophyceae) in two separate clades of the genus Mantoniella. ITS2 synapomorphies support their placement as two new species, Mantoniella beaufortii and Mantoniella baffinensis. Both species have round green cells with diameter between 3 and 5 μm, one long flagellum and a short flagellum (~1 μm) and are covered by spiderweb-like scales, making both species similar to other Mantoniella species. Morphologically, M. beaufortii and M. baffinensis are most similar to the cosmopolitan M. squamata with only minor differences in scale structure distinguishing them. Screening of global marine metabarcoding data sets indicates M. beaufortii has only been recorded in seawater and sea ice samples from the Arctic, while no environmental barcode matches M. baffinensis. Like other Mamiellophyceae genera that have distinct polar and temperate species, the polar distribution of these new species suggests they are cold or ice-adapted Mantoniella species.     

Urban habitat restoration provides a human health benefit through microbiome rewilding: the Microbiome Rewilding Hypothesis

Restoration aims to return ecosystem services, including the human health benefits of exposure to green space. The loss of such exposure with urbanization and industrialization has arguably contributed to an increase in human immune dysregulation. The Biodiversity and Old Friends hypotheses have described the possible mechanisms of this relationship, and suggest that reduced exposure to diverse, beneficial microorganisms can result in negative health consequences. However, it is unclear whether restoration of biodiverse habitat can reverse this effect, and what role the environmental microbiome might have in such recovery. Here, we propose the Microbiome Rewilding Hypothesis, which specifically outlines that restoring biodiverse habitats in urban green spaces can rewild the environmental microbiome to a state that enhances primary prevention of human disease. We support our hypothesis with examples from allied fields, including a case study of active restoration that reversed the degradation of the soil bacterial microbiome of a former pasture. This case study used high‐throughput amplicon sequencing of environmental DNA to assess the quality of a restoration intervention in restoring the soil bacterial microbiome. The method is rapid, scalable, and standardizable, and has great potential as a monitoring tool to assess functional outcomes of green‐space restoration. Evidence for the Microbiome Rewilding Hypothesis will help motivate health professionals, urban planners, and restoration practitioners to collaborate and achieve co‐benefits. Co‐benefits include improved human health outcomes and investment opportunities for biodiversity conservation and restoration.     

Patterns of temporal and enemy niche use by a community of leaf cone moths (Caloptilia) coexisting on maples (Acer) as revealed by metabarcoding

The diversity of herbivorous insects is often considered a function of host plant diversity. However, recent research has uncovered many examples of closely related herbivores using the same host plant(s), suggesting that partitioning of host plants is not the only mechanism generating diversity. Herbivores sharing hosts may utilize different parts of the same plant, but such resource partitioning is often not apparent; hence, the factors that allow closely related herbivores to coexist are still largely undetermined. We examined whether partitioning of phenology or natural enemies may explain the coexistence of leaf cone moths (Caloptilia; Gracillariidae) associated with maples (Acer; Sapindaceae). Larval activity of 10 sympatric Caloptilia species found on nine maple species was monitored every 2–3 weeks for a total of 13 sampling events, and an exhaustive search for internal parasitoid wasps was conducted using high‐throughput sequencing. Blocking primers were used to facilitate the detection of wasp larvae inside moth tissue. We found considerable phenological overlap among Caloptilia species, with two clear peaks in July and September–October. Coexisting Caloptilia species also had largely overlapping parasitoid communities; a total of 13 chalcid and ichneumon wasp species attacked Caloptilia in a nonspecific fashion at an overall parasitism rate of 46.4%. Although coexistence may be facilitated by factors not accounted for in this study, it appears that niche partitioning is not necessary for closely related herbivores to stably coexist on shared hosts. Co‐occurrence without resource partitioning may provide an additional axis along which herbivorous insects attain increased species richness.     

A metabarcoding comparison of windward and leeward airborne algal diversity across the Ko‘olau mountain range on the island of O'ahu,Hawai‘i1

Airborne algae from sites on the windward (n = 3) and leeward (n = 3) sides of the Ko‘olau Mountain range of O‘ahu, Hawai‘i, were sampled for a 16 d period during January and February 2015 using passive collection devices and were characterized using Illumina MiSeq sequencing of the universal plastid amplicon marker. Amplicons were assigned to 3,023 operational taxonomic units (OTUs), which included 1,189 cyanobacteria, 1,009 heterotrophic bacteria, and 304 Eukaryota (of which 284 were algae and land plants). Analyses demonstrated substantially more OTUs at windward than leeward O‘ahu sites during the sampling period. Removal of nonalgal OTUs revealed a greater number of algal reads recovered from windward (839,853) than leeward sites (355,387), with the majority of these being cyanobacteria. The 1,234 total algal OTUs included cyanobacteria, diatoms, cryptophytes, brown algae, chlorophyte green algae, and charophyte green algae. A total of 208 algal OTUs were identified from leeward side samplers (including OTUs in common among samplers) and 1,995 algal OTUs were identified from windward samplers. Barcoding analyses of the most abundant algal OTUs indicated that very few were shared between the windward and leeward sides of the Ko‘olau Mountains, highlighting the localized scale at which these airborne algae communities differ. Back trajectories of air masses arriving on O‘ahu during the sampling period were calculated using the NOAA HY‐SPLIT model and suggested that the sampling period was composed of three large‐scale meteorological events, indicating a diversity of potential sources of airborne algae outside of the Hawaiian Islands.