A validation scale to determine the readiness of environmental DNA assays for routine species monitoring

The use of environmental DNA (eDNA) analysis for species monitoring requires rigorous validation - from field sampling to the analysis of PCR-based results - for meaningful application and interpretation. Assays targeting eDNA released by individual species are typically validated with no predefined criteria to answer specific research questions in one ecosystem. Hence, the general applicability of assays as well as associated uncertainties and limitations, often remain undetermined. The absence of clear guidelines for assay validation prevents targeted eDNA assays from being incorporated into species monitoring and policy; thus, their establishment is essential for realizing the potential of eDNA-based surveys. We describe the measures and tests necessary for successful validation of targeted eDNA assays and the associated pitfalls to form the basis of guidelines. A list of 122 variables was compiled, consolidated into 14 thematic blocks, (e.g. “ in silico analysis”), and arranged on a 5-level validation scale from “incomplete” to “operational” with defined minimum validation criteria for each level. These variables were evaluated for 546 published single-species assays. The resulting dataset was used to provide an overview of current validation practices and test the applicability of the validation scale for future assay rating. Of the 122 variables, 20% to 76% were reported; the majority (30%) of investigated assays were classified as Level 1 (incomplete), and 15% did not achieve this first level. These assays were characterised by minimal in silico and in vitro testing, but their share in annually published eDNA assays has declined since 2014. The meta-analysis demonstrates the suitability of the 5-level validation scale for assessing targeted eDNA assays. It is a user-friendly tool to evaluate previously published assays for future research and routine monitoring, while also enabling the appropriate interpretation of results. Finally, it provides guidance on validation and reporting standards for newly developed assays

Rapid Plant Identification Using Species- and Group-Specific Primers Targeting Chloroplast DNA

Plant identification is challenging when no morphologically assignable parts are available. There is a lack of broadly applicable methods for identifying plants in this situation, for example when roots grow in mixture and for decayed or semi-digested plant material. These difficulties have also impeded the progress made in ecological disciplines such as soil- and trophic ecology. Here, a PCR-based approach is presented which allows identifying a variety of plant taxa commonly occurring in Central European agricultural land. Based on the trnT-F cpDNA region, PCR assays were developed to identify two plant families (Poaceae and Apiaceae), the genera Trifolium and Plantago, and nine plant species: Achillea millefolium, Fagopyrum esculentum, Lolium perenne, Lupinus angustifolius, Phaseolus coccineus, Sinapis alba, Taraxacum officinale, Triticum aestivum, and Zea mays. These assays allowed identification of plants based on size-specific amplicons ranging from 116 bp to 381 bp. Their specificity and sensitivity was consistently high, enabling the detection of small amounts of plant DNA, for example, in decaying plant material and in the intestine or faeces of herbivores. To increase the efficacy of identifying plant species from large number of samples, specific primers were combined in multiplex PCRs, allowing screening for multiple species within a single reaction. The molecular assays outlined here will be applicable manifold, such as for root- and leaf litter identification, botanical trace evidence, and the analysis of herbivory

Monitoring spawning migrations of potamodromous fish species via eDNA

Potamodromous fish are considered important indicators of habitat connectivity in freshwater ecosystems, but they are globally threatened by anthropogenic impacts. Hence, non-invasive techniques are necessary for monitoring during spawning migrations. The use of environmental DNA (eDNA) potentially facilitates these efforts, albeit quantitative examinations of spawning migrations remain so far mostly uncharted. Here, we investigated spawning migrations of Danube bleak, Alburnus mento, and Vimba bream, Vimba vimba, and found a strong correlation between daily visual fish counts and downstream eDNA signals obtained from filtered water samples analysed with digital PCR and end-point PCR coupled with capillary electrophoresis. By accounting for daily discharge fluctuations, it was possible to predict eDNA signal strength from the number of migrating fish: first, the whole spawning reach was taken into account. Second, the model was validated using eDNA signals and fish counts obtained from the upper half of the examined river stretch. Consequently, fish counts and their day-to-day changes could be described via an eDNA-based time series model for the whole migration period. Our findings highlight the capability of eDNA beyond delivering simple presence/absence data towards efficient and informative monitoring of highly dynamic aquatic processes such as spawning migrations of potamodromous fish species.(VLID)4573222Version of recor

Structural and Catalytic Properties of Ag- and Co3O4-Impregnated Strontium Titanium Ferrite SrTi0.7Fe0.3O3-delta in Methanol Steam Reforming

The catalytic performance of Ag- and Co3O4-impregnated strontium titanium ferrite (SrTi0.7Fe0.3O3-delta, STF) has been assessed in methanol steam reforming as a test reaction and accordingly correlated with spectroscopic and structural characterization after each step of a catalytic cycle including preoxidation and prereduction. After prereduction in hydrogen at 400 C, metallic silver and oxidized Co in the form of Co3O4 are present. As for catalysis, it was observed that both Ag and Co3O4 strongly promote the methanol chemistry with respect to pure STF already at low temperatures (230 C for both materials). As a consequence, the selectivity toward carbon monoxide is strongly enhanced in comparison to pure STF. On Co3O4-STF, the catalytic profile is basically dominated by a temperature-dependent complex interplay between methanol steam reforming, methanol dehydrogenation, and the water gas shift equilibrium. The results also prove that despite the obvious inadequacy of H2-prereduced Ag-STF to act as a highly CO2-selective steam reforming catalyst, a potential use as a low-temperature fuel cell anode material might be envisioned. It is shown that Ag effectively lowers the activation barrier for the total oxidation of methanol to CO2, if the reaction is started from the fully preoxidized catalyst. This is explained by fast and efficient supply of lattice oxygen from STF toward Ag via the perovskite-metal phase boundary, which is expected to prevail if Ag-STF is used as a solid oxide fuel cell anode material with continuous electrochemical supply of lattice oxygen.(VLID)2581050Accepted versio

Rhodium-Catalyzed Methanation and Methane Steam Reforming Reactions on Rhodium-Perovskite Systems: Metal-Support Interaction

Metal-support interaction in rhodium-perovskite systems was studied using LSF (La0.6Sr0.4FeO3-[delta]) and STF (SrTi0.7Fe0.3O3-[delta]) supports to disentangle different manifestations of strong or reactive metalsupport interaction. Electron microscopy and catalytic characterization in methane steam reforming/CO2 methanation reveal that reduction in hydrogen at 673K and 873K causes different extents of Fe exsolution. Depending on the perovskite reducibility, Fe-Rh alloy particles are observed. No signs of strong metal-support interaction (i.e., encapsulation of metal particles) by reduced oxide species were observed. As reoxidation in oxygen at 873K did not fully restore the initial structures, the interaction between Rh and the perovskites manifests itself in irreversible alloy formation. Catalytic effects are the suppression of methane reactivity with increasing prereduction temperature. The results show the limits of the strong metal-support interaction concept in complex metal-oxide systems.(VLID)1371554Accepted versio