Chronic inflammation decreases HSC fitness by activating the druggable Jak/Stat3 signaling pathway.

Chronic inflammation represents a major threat to human health since long-term systemic inflammation is known to affect distinct tissues and organs. Recently, solid evidence demonstrated that chronic inflammation affects hematopoiesis; however, how chronic inflammation affects hematopoietic stem cells (HSCs) on the mechanistic level is poorly understood. Here, we employ a mouse model of chronic multifocal osteomyelitis (CMO) to assess the effects of a spontaneously developed inflammatory condition on HSCs. We demonstrate that hematopoietic and nonhematopoietic compartments in CMO BM contribute to HSC expansion and impair their function. Remarkably, our results suggest that the typical features of murine multifocal osteomyelitis and the HSC phenotype are mechanistically decoupled. We show that the CMO environment imprints a myeloid gene signature and imposes a pro-inflammatory profile on HSCs. We identify IL-6 and the Jak/Stat3 signaling pathway as critical mediators. However, while IL-6 and Stat3 blockage reduce HSC numbers in CMO mice, only inhibition of Stat3 activity significantly rescues their fitness. Our data emphasize the detrimental effects of chronic inflammation on stem cell function, opening new venues for treatment.

Trait-mediated responses of caterpillar communities to spongy moth outbreaks and subsequent tebufenozide treatments.

Outbreaks of the spongy moth Lymantria dispar can have devastating impacts on forest resources and ecosystems. Lepidoptera-specific insecticides, such as Bacillus thuringiensis var. kurstaki (BTK) and tebufenozide, are often deployed to prevent heavy defoliation of the forest canopy. While it has been suggested that using BTK poses less risk to non-target Lepidoptera than leaving an outbreak untreated, in situ testing of this assumption has been impeded by methodological challenges. The trade-offs between insecticide use and outbreaks have yet to be addressed for tebufenozide, which is believed to have stronger side effects than BTK. We investigated the short-term trade-offs between tebufenozide treatments and no-action strategies for the non-target herbivore community in forest canopies. Over 3 years, Lepidoptera and Symphyta larvae were sampled by canopy fogging in 48 oak stands in southeast Germany during and after a spongy moth outbreak. Half of the sites were treated with tebufenozide and changes in canopy cover were monitored. We contrasted the impacts of tebufenozide and defoliator outbreaks on the abundance, diversity, and functional structure of chewing herbivore communities. Tebufenozide treatments strongly reduced Lepidoptera up to 6 weeks after spraying. Populations gradually converged back to control levels after 2 years. Shelter-building species dominated caterpillar assemblages in treated plots in the post-spray weeks, while flight-dimorphic species were slow to recover and remained underrepresented in treated stands 2 years post-treatment. Spongy moth outbreaks had minor effects on leaf chewer communities. Summer Lepidoptera decreased only when severe defoliation occurred, whereas Symphyta declined 1 year after defoliation. Polyphagous species with only partial host plant overlap with the spongy moth were absent from heavily defoliated sites, suggesting greater sensitivity of generalists to defoliation-induced plant responses. These results demonstrate that both tebufenozide treatments and spongy moth outbreaks alter canopy herbivore communities. Tebufenozide had a stronger and longer lasting impact, but it was restricted to Lepidoptera, whereas the outbreak affected both Lepidoptera and Symphyta. These results are tied to the fact that only half of the outbreak sites experienced severe defoliation. This highlights the limited accuracy of current defoliation forecast methods, which are used as the basis for the decision to spray insecticides.

The structure of tropical bat-plant interaction networks during an extreme El Niño-Southern Oscillation event.

Interaction network structure reflects the ecological mechanisms acting within biological communities, which are affected by environmental conditions. In tropical forests, higher precipitation usually increases fruit production, which may lead frugivores to increase specialization, resulting in more modular and less nested animal-plant networks. In these ecosystems, El Niño is a major driver of precipitation, but we still lack knowledge of how species interactions change under this influence. To understand bat-plant network structure during an extreme El Niño-Southern Oscillation event, we determined the links between plantivorous bat species and the plants they consume by DNA barcoding seeds and pulp in bat faeces. These interactions were recorded in the dry forest and rainforest of Costa Rica, during the dry and the wet seasons of an extreme El Niño year. From these we constructed seasonal and whole-year bat-plant networks and analysed their structures and dissimilarities. In general, networks had low nestedness, had high modularity, and were dominated by one large compartment which included most species and interactions. Contrary to our expectations, networks were less nested and more modular in drier conditions, both in the comparison between forest types and between seasons. We suggest that increased competition, when resources are scarce during drier seasons and habitats, lead to higher resource partitioning among bats and thus higher modularity. Moreover, we have found similar network structures between dry and rainforests during El Niño and non-El Niño years. Finally, most interaction dissimilarity among networks occurred due to interaction rewiring among species, potentially driven by seasonal changes in resource availability.

DNA metabarcoding illuminates dietary niche partitioning by African large herbivores.

Niche partitioning facilitates species coexistence in a world of limited resources, thereby enriching biodiversity. For decades, biologists have sought to understand how diverse assemblages of large mammalian herbivores (LMH) partition food resources. Several complementary mechanisms have been identified, including differential consumption of grasses versus nongrasses and spatiotemporal stratification in use of different parts of the same plant. However, the extent to which LMH partition food-plant species is largely unknown because comprehensive species-level identification is prohibitively difficult with traditional methods. We used DNA metabarcoding to quantify diet breadth, composition, and overlap for seven abundant LMH species (six wild, one domestic) in semiarid African savanna. These species ranged from almost-exclusive grazers to almost-exclusive browsers: Grass consumption inferred from mean sequence relative read abundance (RRA) ranged from >99% (plains zebra) to <1% (dik-dik). Grass RRA was highly correlated with isotopic estimates of % grass consumption, indicating that RRA conveys reliable quantitative information about consumption. Dietary overlap was greatest between species that were similar in body size and proportional grass consumption. Nonetheless, diet composition differed between all species-even pairs of grazers matched in size, digestive physiology, and location-and dietary similarity was sometimes greater across grazing and browsing guilds than within them. Such taxonomically fine-grained diet partitioning suggests that coarse trophic categorizations may generate misleading conclusions about competition and coexistence in LMH assemblages, and that LMH diversity may be more tightly linked to plant diversity than is currently recognized.

Phylogeny, character evolution, and biogeography of Cuscuta (dodders; Convolvulaceae) inferred from coding plastid and nuclear sequences.

PREMISE OF THE STUDY: The parasitic genus Cuscuta, containing some 200 species circumscribed traditionally in three subgenera, is nearly cosmopolitan, occurring in a wide range of habitats and hosts. Previous molecular studies, on subgenera Grammica and Cuscuta, delimited major clades within these groups. However, the sequences used were unalignable among subgenera, preventing the phylogenetic comparison across the genus. METHODS: We conducted a broad phylogenetic study using rbcL and nrLSU sequences covering the morphological, physiological, and geographical diversity of Cuscuta. We used parsimony methods to reconstruct ancestral states for taxonomically important characters. Biogeographical inferences were obtained using statistical and Bayesian approaches. KEY RESULTS: Four well-supported major clades are resolved. Two of them correspond to subgenera Monogynella and Grammica. Subgenus Cuscuta is paraphyletic, with section Pachystigma sister to subgenus Grammica. Previously described cases of strongly supported discordance between plastid and nuclear phylogenies, interpreted as reticulation events, are confirmed here and three new cases are detected. Dehiscent fruits and globose stigmas are inferred as ancestral character states, whereas the ancestral style number is ambiguous. Biogeographical reconstructions suggest an Old World origin for the genus and subsequent spread to the Americas as a consequence of one long-distance dispersal. CONCLUSIONS: Hybridization may play an important yet underestimated role in the evolution of Cuscuta. Our results disagree with scenarios of evolution (polarity) previously proposed for several taxonomically important morphological characters, and with their usage and significance. While several cases of long-distance dispersal are inferred, vicariance or dispersal to adjacent areas emerges as the dominant biogeographical pattern.

DNA barcoding aids in generating a preliminary checklist of the lichens and allied fungi of Calvert Island, British Columbia: Results from the 2018 Hakai Terrestrial BioBlitz.

Background: Bioblitzes are a tool for the rapid appraisal of biodiversity and are particularly useful in remote and understudied regions and for understudied taxa. Lichens are an example of an often overlooked group, despite being widespread in virtually all terrestrial ecosystems and having many important ecological functions. New information: We report the lichens and allied fungi collected during the 2018 terrestrial bioblitz conducted on Calvert Island on the Central Coast of British Columbia, Canada. We identified 449 specimens belonging to 189 species in 85 genera, increasing the total number of species known from Calvert Island to 194, and generated Internal Transcribed Spacer (ITS) sequences for 215 specimens from 121 species. Bryoriafurcellata, Chaenothecopsislecanactidis and C.nigripunctata were collected for the first time in British Columbia. We also found Pseudocyphellariarainierensis, which is listed as Special Concern on the federal Species at Risk Act, and other rarely reported species in British Columbia including Opegraphasphaerophoricola, Protomicarealimosa, Raesaeneniahuuskonenii and Sareadifformis. We demonstrate that DNA barcoding improves the scope and accuracy of expert-led bioblitzes by facilitating the detection of cryptic species and allowing for consistent identification of chemically and morphologically overlapping taxa. Despite the spatial and temporal limitations of our study, the results highlight the value of intact forest ecosystems on the Central Coast of British Columbia for lichen biodiversity, education and conservation.

Cenozoic extinction and recolonization in the New Zealand flora: the case of the fleshy-fruited epacrids (Styphelieae, Styphelioideae, Ericaceae).

The origins and evolutionary history of the New Zealand flora has been the subject of much debate. The recent description of Cyathodophyllum novaezelandieae from early Miocene sediments in New Zealand provides possible evidence for the antiquity of the fleshy fruited epacrids (tribe Styphelieae, Ericaceae) in New Zealand. Yet the extant species in this tribe are thought to be very closely related to or conspecific with Australian taxa, suggesting recent trans-Tasman origins. In order to investigate the origins and evolution of the extant New Zealand Styphelieae we produced molecular phylogenetic trees based on sequences of three plastid regions that include representatives of all the genera of the tribe and eight of the ten New Zealand species. We estimated the range of minimum ages of the New Zealand lineages with Bayesian relaxed-clock analyses using different calibration methods and relative dating. We found strong support for each of the eight extant species of New Zealand Styphelieae being a distinct lineage that is nested within an Australian clade. In all except one case the sister is from Tasmania and/or the east coast of mainland Australia; for Acrothamnus colensoi the sister is in New Guinea. Estimated dates indicate that all of the New Zealand lineages diverged from their non-New Zealand sisters within the last 7 Ma. Time discontinuity between the fossil C.novae-zelandiae (20-23 Ma) and the origins of the extant New Zealand lineages (none older than 5 Ma) indicates that the fossil and extant Styphelieae in New Zealand are not related. The relative dating analysis showed that to accept this relationship, it would be necessary to accept that the Styphelieae arose in the early-mid Mesozoic (210-120 Ma), which is starkly at odds with multiple lines of evidence on the age of Ericales and indeed the angiosperms. Therefore, our results do not support the hypothesis that Styphelieae have been continuously present in New Zealand since the early Miocene. Instead they suggest a historical biogeographical scenario in which the lineage to which C. novae-zelandiae belongs went extinct in New Zealand, and the extant New Zealand Styphelieae are derived from Australian lineages that recolonised (presumably by long distance dispersal) no earlier than the late Miocene to Pliocene.

Plastid genome evolution across the genus Cuscuta (Convolvulaceae): two clades within subgenus Grammica exhibit extensive gene loss.

The genus Cuscuta (Convolvulaceae, the morning glory family) is one of the most intensely studied lineages of parasitic plants. Whole plastome sequencing of four Cuscuta species has demonstrated changes to both plastid gene content and structure. The presence of photosynthetic genes under purifying selection indicates that Cuscuta is cryptically photosynthetic. However, the tempo and mode of plastid genome evolution across the diversity of this group (~200 species) remain largely unknown. A comparative investigation of plastid genome content, grounded within a phylogenetic framework, was conducted using a slot-blot Southern hybridization approach. Cuscuta was extensively sampled (~56% of species), including groups previously suggested to possess more altered plastomes compared with other members of this genus. A total of 56 probes derived from all categories of protein-coding genes, typically found within the plastomes of flowering plants, were used. The results indicate that two clades within subgenus Grammica (clades 'O' and 'K') exhibit substantially more plastid gene loss relative to other members of Cuscuta. All surveyed members of the 'O' clade show extensive losses of plastid genes from every category of genes typically found in the plastome, including otherwise highly conserved small and large ribosomal subunits. The extent of plastid gene losses within this clade is similar in magnitude to that observed previously in some non-asterid holoparasites, in which the very presence of a plastome has been questioned. The 'K' clade also exhibits considerable loss of plastid genes. Unlike in the 'O' clade, in which all species seem to be affected, the losses in clade 'K' progress phylogenetically, following a pattern consistent with the Evolutionary Transition Series hypothesis. This clade presents an ideal opportunity to study the reduction of the plastome of parasites 'in action'. The widespread plastid gene loss in these two clades is hypothesized to be a consequence of the complete loss of photosynthesis. Additionally, taxa that would be the best candidates for entire plastome sequencing are identified in order to investigate further the loss of photosynthesis and reduction of the plastome within Cuscuta.

Local eukaryotic and bacterial stream community assembly is shaped by regional land use effects.

With anticipated expansion of agricultural areas for food production and increasing intensity of pressures stemming from land-use, it is critical to better understand how species respond to land-use change. This is particularly true for microbial communities which provide key ecosystem functions and display fastest responses to environmental change. However, regional land-use effects on local environmental conditions are often neglected, and, hence, underestimated when investigating community responses. Here we show that the effects stemming from agricultural and forested land use are strongest reflected in water conductivity, pH and phosphorus concentration, shaping microbial communities and their assembly processes. Using a joint species distribution modelling framework with community data based on metabarcoding, we quantify the contribution of land-use types in determining local environmental variables and uncover the impact of both, land-use, and local environment, on microbial stream communities. We found that community assembly is closely linked to land-use type but that the local environment strongly mediates the effects of land-use, resulting in systematic variation of taxon responses to environmental conditions, depending on their domain (bacteria vs. eukaryote) and trophic mode (autotrophy vs. heterotrophy). Given that regional land-use type strongly shapes local environments, it is paramount to consider its key role in shaping local stream communities.

Identification of the vascular plants of Churchill, Manitoba, using a DNA barcode library.

BACKGROUND: Because arctic plant communities are highly vulnerable to climate change, shifts in their composition require rapid, accurate identifications, often for specimens that lack diagnostic floral characters. The present study examines the role that DNA barcoding can play in aiding floristic evaluations in the arctic by testing the effectiveness of the core plant barcode regions (rbcL, matK) and a supplemental ribosomal DNA (ITS2) marker for a well-studied flora near Churchill, Manitoba. RESULTS: This investigation examined 900 specimens representing 312 of the 354 species of vascular plants known from Churchill. Sequencing success was high for rbcL: 95% for fresh specimens and 85% for herbarium samples (mean age 20 years). ITS2 worked equally well for the fresh and herbarium material (89% and 88%). However, sequencing success was lower for matK, despite two rounds of PCR amplification, which reflected less effective primer binding and sensitivity to the DNA degradation (76% of fresh, 45% of herbaria samples). A species was considered as taxonomically resolved if its members showed at least one diagnostic difference from any other taxon in the study and formed a monophyletic clade. The highest species resolution (69%) was obtained by combining information from all three genes. The joint sequence information for rbcL and matK distinguished 54% of 286 species, while rbcL and ITS2 distinguished 63% of 285 species. Discrimination of species within Salix, which constituted 8% of the flora, was particularly problematic. Despite incomplete resolution, the barcode results revealed 22 misidentified herbarium specimens, and enabled the identification of field specimens which were otherwise too immature to identify. Although seven cases of ITS2 paralogy were noted in the families Cyperaceae, Juncaceae and Juncaginaceae, this intergenic spacer played an important role in resolving congeneric plant species at Churchill. CONCLUSIONS: Our results provided fast and cost-effective solution to create a comprehensive, effective DNA barcode reference library for a local flora.

Futurization of Aging: Subjective Beliefs and Effects.

Aging in the face of an increasing population and growing life expectancy is considered one of the major demographic challenges in modern society. Previous research has revealed that quality of life in aging could significantly differ depending on the resources one possesses. However, little attention has been given to the mechanisms of formation of these resources and the role of intentionality. In the present study, we identified 22 strategies that favor a better life quality in aging and analyzed them from the perspective of subjective beliefs and reported performance. Our sample was adults (n = 72) aged 57-65, living in St. Petersburg, Russia. The results showed that although participants were aware of the strategies that favor aging, their reported performance ranged on a scale from average to infrequent use of these strategies. We found that subjective beliefs about the role of psychological resources for better aging predicted higher scores on subjective beliefs about the role of lifestyle resources and the reported performance of psychological resources. Our results suggest that there is a gap between subjective beliefs about the controllability of aging processes and the transformation of these beliefs into real performance.

Improved hematopoietic stem cell transplantation upon inhibition of natural killer cell-derived interferon-gamma.

Hematopoietic stem cell transplantation (HSCT) is a frequent therapeutic approach to restore hematopoiesis in patients with hematologic diseases. Patients receive a hematopoietic stem cell (HSC)-enriched donor cell infusion also containing immune cells, which may have a beneficial effect by eliminating residual neoplastic cells. However, the effect that donor innate immune cells may have on the donor HSCs has not been deeply explored. Here, we evaluate the influence of donor natural killer (NK) cells on HSC fate, concluded that NK cells negatively affect HSC frequency and function, and identified interferon-gamma (IFNγ) as a potential mediator. Interestingly, improved HSC fitness was achieved by NK cell depletion from murine and human donor infusions or by blocking IFNγ activity. Thus, our data suggest that suppression of inflammatory signals generated by donor innate immune cells can enhance engraftment and hematopoietic reconstitution during HSCT, which is particularly critical when limited HSC numbers are available and the risk of engraftment failure is high.

Loss of all plastid ndh genes in Gnetales and conifers: extent and evolutionary significance for the seed plant phylogeny.

The exact phylogenetic position of Gnetales, a small, highly modified group of gymnosperms with an accelerated rate of molecular evolution, is one of the most challenging issues for seed plant systematics. Recent results from entire plastid genome (ptDNA) sequencing revealed the absence of the entire suite of plastid ndh genes in several species of Gnetales and the pine family (Pinaceae) potentially highlighting a major structural feature linking these two groups-concerted loss of all plastid genes for the NADH dehydrogenase complex. However, the precise extent of ndh gene loss in gymnosperms has not been surveyed. Using a slot-blot hybridization method, we probed all 11 ndh genes in 162 species from 70 of 85 gymnosperm genera. We find that all ndh genes are absent across Gnetales and Pinaceae, but not in any other group of gymnosperms. This feature represents either a major synapomorphy for a clade consisting of these two lineages or, less likely, a convergent loss. Our survey substantially extends previous inferences based on more limited sampling and, if the former evolutionary interpretation is correct, it provides additional support for the contentious "gnepine" hypothesis, which places Gnetales as sister to Pinaceae.

Features of diclofenac biodegradation by Rhodococcus ruber IEGM 346.

This study investigated the ability of rhodococci to biodegrade diclofenac (DCF), one of the polycyclic non-steroidal anti-inflammatory drugs (NSAIDs) most frequently detected in the environment. Rhodococcus ruber strain IEGM 346 capable of complete DCF biodegradation (50 µg/L) over 6 days was selected. It is distinguished by the ability to degrade DCF at high (50 mg/L) concentrations unlike other known biodegraders. The DCF decomposition process was accelerated by adding glucose and due to short-term cell adaptation to 5 µg/L DCF. The most typical responses to DCF exposure observed were the changed ζ-potential of bacterial cells; increased cell hydrophobicity and total cell lipid content; multi-cellular conglomerates formed; and the changed surface-to-volume ratio. The obtained findings are considered as mechanisms of rhodococcal adaptation and hence their increased resistance to toxic effects of this pharmaceutical pollutant. The proposed pathways of bacterial DCF metabolisation were described. The data confirming the C-N bond cleavage and aromatic ring opening in the DCF structure were obtained.

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.

Finding the pond through the weeds: eDNA reveals underestimated diversity of pondweeds.

PREMISE OF THE STUDY: The detection of environmental DNA (eDNA) using high-throughput sequencing has rapidly emerged as a method to detect organisms from environmental samples. However, eDNA studies of aquatic biomes have focused on surveillance of animal species with less emphasis on plants. Pondweeds are important bioindicators of freshwater ecosystems, although their diversity is underestimated due to difficulties in morphological identification and monitoring. METHODS: A protocol was developed to detect pondweeds in water samples using atpB-rbcL and ITS2 markers. The water samples were collected from the Grand River within the rare Charitable Research Reserve, Ontario (RARE). Short fragments were amplified using primers targeting pondweeds, sequenced on an Ion Torrent Personal Genome Machine, and assigned to the taxonomy using a local DNA reference library and GenBank. RESULTS: We detected two species earlier documented at the experimental site during ecological surveys (Potamogeton crispus and Stuckenia pectinata) and three species new to the RARE checklist (P. foliosus, S. filiformis, and Zannichellia palustris). DISCUSSION: Our targeted approach to track the species composition of pondweeds in freshwater ecosystems revealed underestimation of their diversity. This result suggests that eDNA is an effective tool for monitoring plant diversity in aquatic habitats.

Testing the Efficacy of DNA Barcodes for Identifying the Vascular Plants of Canada.

Their relatively slow rates of molecular evolution, as well as frequent exposure to hybridization and introgression, often make it difficult to discriminate species of vascular plants with the standard barcode markers (rbcL, matK, ITS2). Previous studies have examined these constraints in narrow geographic or taxonomic contexts, but the present investigation expands analysis to consider the performance of these gene regions in discriminating the species in local floras at sites across Canada. To test identification success, we employed a DNA barcode reference library with sequence records for 96% of the 5108 vascular plant species known from Canada, but coverage varied from 94% for rbcL to 60% for ITS2 and 39% for matK. Using plant lists from 27 national parks and one scientific reserve, we tested the efficacy of DNA barcodes in identifying the plants in simulated species assemblages from six biogeographic regions of Canada using BLAST and mothur. Mean pairwise distance (MPD) and mean nearest taxon distance (MNTD) were strong predictors of barcode performance for different plant families and genera, and both metrics supported ITS2 as possessing the highest genetic diversity. All three genes performed strongly in assigning the taxa present in local floras to the correct genus with values ranging from 91% for rbcL to 97% for ITS2 and 98% for matK. However, matK delivered the highest species discrimination (~81%) followed by ITS2 (~72%) and rbcL (~44%). Despite the low number of plant taxa in the Canadian Arctic, DNA barcodes had the least success in discriminating species from this biogeographic region with resolution ranging from 36% with rbcL to 69% with matK. Species resolution was higher in the other settings, peaking in the Woodland region at 52% for rbcL and 87% for matK. Our results indicate that DNA barcoding is very effective in identifying Canadian plants to a genus, and that it performs well in discriminating species in regions where floristic diversity is highest.

Using herbarium-derived DNAs to assemble a large-scale DNA barcode library for the vascular plants of Canada.

PREMISE OF THE STUDY: Constructing complete, accurate plant DNA barcode reference libraries can be logistically challenging for large-scale floras. Here we demonstrate the promise and challenges of using herbarium collections for building a DNA barcode reference library for the vascular plant flora of Canada. METHODS: Our study examined 20,816 specimens representing 5076 of 5190 vascular plant species in Canada (98%). For 98% of the specimens, at least one of the DNA barcode regions was recovered from the plastid loci rbcL and matK and from the nuclear ITS2 region. We used beta regression to quantify the effects of age, type of preservation, and taxonomic affiliation (family) on DNA sequence recovery. RESULTS: Specimen age and method of preservation had significant effects on sequence recovery for all markers, but influenced some families more (e.g., Boraginaceae) than others (e.g., Asteraceae). DISCUSSION: Our DNA barcode library represents an unparalleled resource for metagenomic and ecological genetic research working on temperate and arctic biomes. An observed decline in sequence recovery with specimen age may be associated with poor primer matches, intragenomic variation (for ITS2), or inhibitory secondary compounds in some taxa.

Correction: Authentication of Herbal Supplements Using Next-Generation Sequencing.

[This corrects the article DOI: 10.1371/journal.pone.0156426.].

Authentication of Herbal Supplements Using Next-Generation Sequencing.

BACKGROUND: DNA-based testing has been gaining acceptance as a tool for authentication of a wide range of food products; however, its applicability for testing of herbal supplements remains contentious. METHODS: We utilized Sanger and Next-Generation Sequencing (NGS) for taxonomic authentication of fifteen herbal supplements representing three different producers from five medicinal plants: Echinacea purpurea, Valeriana officinalis, Ginkgo biloba, Hypericum perforatum and Trigonella foenum-graecum. Experimental design included three modifications of DNA extraction, two lysate dilutions, Internal Amplification Control, and multiple negative controls to exclude background contamination. Ginkgo supplements were also analyzed using HPLC-MS for the presence of active medicinal components. RESULTS: All supplements yielded DNA from multiple species, rendering Sanger sequencing results for rbcL and ITS2 regions either uninterpretable or non-reproducible between the experimental replicates. Overall, DNA from the manufacturer-listed medicinal plants was successfully detected in seven out of eight dry herb form supplements; however, low or poor DNA recovery due to degradation was observed in most plant extracts (none detected by Sanger; three out of seven-by NGS). NGS also revealed a diverse community of fungi, known to be associated with live plant material and/or the fermentation process used in the production of plant extracts. HPLC-MS testing demonstrated that Ginkgo supplements with degraded DNA contained ten key medicinal components. CONCLUSION: Quality control of herbal supplements should utilize a synergetic approach targeting both DNA and bioactive components, especially for standardized extracts with degraded DNA. The NGS workflow developed in this study enables reliable detection of plant and fungal DNA and can be utilized by manufacturers for quality assurance of raw plant materials, contamination control during the production process, and the final product. Interpretation of results should involve an interdisciplinary approach taking into account the processes involved in production of herbal supplements, as well as biocomplexity of plant-plant and plant-fungal biological interactions.