scholarly journals An assay validation framework to compare and evaluate targeted environmental DNA assays for routine species monitoring

2021 ◽  
Vol 4 ◽  
Author(s):  
Bettina Thalinger ◽  
Kristy Deiner ◽  
Lynsey Harper ◽  
Helen Rees ◽  
Rosetta Blackman ◽  
...  
Keyword(s):  
Meta Analysis ◽  
Environmental Dna ◽  
Single Species ◽  
End Users ◽  
Specific System ◽  
Initial Development ◽  
Assay Validation ◽  
Non Invasive ◽  
Detection Probabilities ◽  
User Friendly

Environmental DNA (eDNA) analysis utilises trace DNA released by organisms into their environment for species detection and is revolutionising non ‐ invasive species and biodiversity monitoring. However, this technology requires rigorous validation along the whole workflow – from field sampling to statistical analysis – to ensure appropriate and meaningful interpretation of results. Targeted eDNA assays are often validated within a specific system and with particular aims, but without fulfilling predefined criteria. Consequently, their applicability beyond initial development often remains undetermined. Additionally, there tends to be poor understanding of the uncertainties and limitations associated with already published assays and thus potentially inappropriate interpretation of the results they produce. The lack of a “gold standard” limits the incorporation of targeted eDNA assays into species monitoring and policy making by end-users and is therefore key for the future implementation of eDNA-based surveys. Here, we present a framework (https://edna-validation.com/) and user-friendly criteria for the classification of assays, which is based on previous validation efforts. A 5 ‐ level assay validation scale (“incomplete” to “operational”) was defined by reviewing the current eDNA literature and conducting a meta-analysis on sampling, laboratory practices, detection limits, and detection probabilities. The so far published single species eDNA assays were reviewed for their performance in this new framework and we identified steps within the validation process that often remain untouched. Finally, we provide guidance for end ‐ users as to which criteria are most important for validation and suggest how results obtained from assays at different levels of the validation scale should be interpreted.

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

2020 ◽  
Author(s):  
Bettina Thalinger ◽  
Kristy Deiner ◽  
Lynsey R. Harper ◽  
Helen C. Rees ◽  
Rosetta C. Blackman ◽  
...  
Keyword(s):  
In Silico ◽  
Meta Analysis ◽  
Environmental Dna ◽  
Single Species ◽  
Future Research ◽  
Individual Species ◽  
General Applicability ◽  
Routine Monitoring ◽  
User Friendly

AbstractThe use of environmental DNA (eDNA) analysis for species monitoring requires rigorous validation - from field sampling to interpretation 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. Their general applicability, uncertainties and limitations often remain undetermined. The absence of clear guidelines prevents targeted eDNA assays from being incorporated into species monitoring and policy, thus their establishment will be key for the future implementation 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, such as “in silico analysis”, and arranged on a 5-level validation scale from “incomplete” to “operational”. Additionally, minimum validation criteria were defined 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. 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 total number of reported variables ranged from 20% to 76% and deviated both between and within levels. 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 appropriate interpretation of results. Finally, it provides guidance on validation and reporting standards for newly developed assays.

Evaluation of attractants for non-invasive studies of Iberian carnivore communities

2011 ◽  
Vol 38 (5) ◽  
pp. 446 ◽  
Author(s):  
Pedro Monterroso ◽  
Paulo Célio Alves ◽  
Pablo Ferreras
Keyword(s):  
Survey Methods ◽  
Species Abundance ◽  
Field Tests ◽  
Field Evaluation ◽  
Single Species ◽  
Field Studies ◽  
Field Trials ◽  
Non Invasive ◽  
Carnivore Species ◽  
Detection Probabilities

Context The estimation of population parameters for mammalian carnivore species is a challenging task because of their low densities and large home ranges, which make detection probabilities very low. Several factors, such as the species abundance, habitat structure or the use of an attractant affect carnivore detection probabilities; however, attractants are the most easily manipulated. Some previous research suggests that the use of effective attractants can significantly increase detection probabilities. Aims To assess the effectiveness of several attractants for Iberian carnivores, and to evaluate their usefulness for non-invasive survey methods. Methods The responses of seven carnivore species to six potential attractants were evaluated through cafeteria-like experiments with captive specimens. A selectivity index was applied to assess the relative attractiveness of each tested substance. The enclosure tests were followed by field trials with camera-trapping, using the most promising attractants for field evaluation of their efficiency. Key results Enclosure trials revealed that lynx urine was the most effective and generalist attractant because it successfully attracted six of the seven species tested. Rubbing behaviour was also induced in the greatest number of species by lynx urine. Field tests using a combination of lynx urine and valerian extract solution induced investigative behaviours in over 50% of all detection events in all species, with the exception of the Eurasian badger. Conclusions No single attractant is effective for all species. Nevertheless, a combination of lynx urine and valerian solution should efficiently attract the majority of species present in Iberian carnivore communities. Furthermore, some species exhibit a rubbing behaviour when they come in contact with the attractants. Regardless of the generalist efficiency of the lynx urine, other tested substances revealed promising results for single-species monitoring. Implications Our results provide a baseline for selecting attractants in survey and monitoring programs that focus on carnivore species. The rubbing behaviours exhibited by several of the species tested suggest the use of these attractants could improve the efficiency of field studies that rely on rub-pads for the collection of biological samples.

scholarly journals The use of multiple markers and internal positive controls significantly improves species eDNA detection rates and data reliability

2021 ◽  
Vol 4 ◽  
Author(s):  
Rein Brys ◽  
Teun Everts ◽  
David Halfmaerten ◽  
Sabrina Neyrinck ◽  
Quentin Mauvisseau
Keyword(s):  
False Negative ◽  
Strong Support ◽  
Environmental Dna ◽  
Single Species ◽  
Detection Rates ◽  
Multiple Markers ◽  
Molecular Approaches ◽  
Detection Probabilities ◽  
Elusive Species ◽  
Positive Controls

In recent years, environmental DNA analyses became increasingly integrated to detect and monitor the presence and abundance of rare organisms, especially in inaccessible aquatic habitats. Although it is generally proven that detection probabilities of eDNA surveys exceed those obtained via conventional techniques, these molecular approaches are, however, also subjected to detection limitations and levels of uncertainty. Besides improvements that can be made in terms of sampling design, volumes of filtered water, and the effective quantity of DNA that is finally analysed, the sensitivity of eDNA surveys is inherently determined by the number of target eDNA copies suspended in the water column. Here we show that multiplexing different primer/probe assays for the same species, but targeting amplicons situated at different loci, is a surprisingly overlooked aspect that can substantially contribute to reduce these limitations and increase the sensitivity of single-species detections. By empirically testing a large number of natural eDNA samples via ddPCR, we reveal that the use of multiple markers can significantly lower the LOD and LOQ of rare and elusive species, such as the invasive American bullfrog and the endangered European weather loach in a variety of different water bodies, such as ponds, lakes, streams, canals, etc. Especially at very low eDNA concentrations of both target species, our results showed that analysing mulitple loci significantly increased detection probabilities and lowered stochasticity effects, and thus ultimately reduces PCR costs when analysed in multiplex. The validation and use of more than one assay taregtting a single species, may further increase the confidence of positive detections. Finally, we illustrate that the implementation of internal positive controls (IPC's), is an absolute must for accurate validation of eDNA workflows and reliable interpretation of the generated data. IPC’s not only help to track down degraded and inhibited samples, to avoid false-negative detections, it also offers insights into extraction efficiency, indispensable for accurate quantification of population densities. Overall, our findings provide strong support that the multiplexing of multiple markers on different loci in combination with the use of internal positive controls ensures increased detection rates at very low eDNA concentrations and generates more robust and reliable data.

Relationship between total plasma homocysteine and the risk of aneurysms – a meta-analysis

VASA ◽  
2020 ◽  
pp. 1-6
Author(s):  
Hanji Zhang ◽  
Dexin Yin ◽  
Yue Zhao ◽  
Yezhou Li ◽  
Dejiang Yao ◽  
...  
Keyword(s):  
Large Scale ◽  
Meta Analysis ◽  
Single Species ◽  
Total Plasma ◽  
Control Groups ◽  
Randomized Controlled ◽  
Randomized Controlled Studies ◽  
The Difference ◽  
The Relationship ◽  
Healthy Participants

Summary: Our meta-analysis focused on the relationship between homocysteine (Hcy) level and the incidence of aneurysms and looked at the relationship between smoking, hypertension and aneurysms. A systematic literature search of Pubmed, Web of Science, and Embase databases (up to March 31, 2020) resulted in the identification of 19 studies, including 2,629 aneurysm patients and 6,497 healthy participants. Combined analysis of the included studies showed that number of smoking, hypertension and hyperhomocysteinemia (HHcy) in aneurysm patients was higher than that in the control groups, and the total plasma Hcy level in aneurysm patients was also higher. These findings suggest that smoking, hypertension and HHcy may be risk factors for the development and progression of aneurysms. Although the heterogeneity of meta-analysis was significant, it was found that the heterogeneity might come from the difference between race and disease species through subgroup analysis. Large-scale randomized controlled studies of single species and single disease species are needed in the future to supplement the accuracy of the results.

SIGN IN Welcome!Log into your account your username your password Forgot your password Privacy policy PASSWORD RECOVERY Recover your password your email Struktol Home New fast measuring method for process optimization of sucrose crystallization New fast measuring method for process optimization of sucrose crystallization

2020 ◽  
pp. 49-52
Author(s):  
Trine Aabo Andersen
Keyword(s):  
Image Analysis ◽  
Process Optimization ◽  
Crystal Size ◽  
Measuring Method ◽  
Sieve Analysis ◽  
Image Analysis System ◽  
Line System ◽  
Non Invasive ◽  
Analysis System ◽  
User Friendly

A new fast measuring method for process optimization of sucrose crystallization using image analysis based on high quality images and algorithms is introduced. With the mobile, non-invasive at-line system all steps of the sucrose crystallization can be measured to determine the crystal size distribution. The image analysis system is easy to operate and is as well an efficient laboratory solution with user-friendly and customized software. In comparison to sieve analysis, image analyses performed with the ParticleTech Solution have been proven to be reliable.

scholarly journals Hepatitis B Core-Related Antigen as Surrogate Biomarker of Intrahepatic Hepatitis B Virus Covalently-Closed-Circular DNA in Patients with Chronic Hepatitis B: A Meta-Analysis

Diagnostics ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 187
Author(s):  
Gian Paolo Caviglia ◽  
Angelo Armandi ◽  
Chiara Rosso ◽  
Davide Giuseppe Ribaldone ◽  
Rinaldo Pellicano ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hepatitis B Surface Antigen ◽  
Meta Analysis ◽  
Circular Dna ◽  
Non Invasive ◽  
Hbv Cccdna ◽  
B Virus ◽  
Chronic Hbv ◽  
Intrahepatic Hbv

Hepatitis B virus (HBV) covalently-closed-circular (ccc)DNA is the key molecule responsible for viral persistence within infected hepatocytes. The evaluation of HBV cccDNA is crucial for the management of patients with chronic HBV infection and for the personalization of treatment. However, the need for liver biopsy is the principal obstacle for the assessment of intrahepatic HBV cccDNA. In the last decade, several studies have investigated the performance of hepatitis B core-related antigen (HBcrAg) as a surrogate of HBV cccDNA amount in the liver. In this meta-analysis, we collected 14 studies (1271 patients) investigating the correlation between serum HBcrAg and intrahepatic HBV cccDNA. Serum HBcrAg showed a high correlation with intrahepatic HBV cccDNA (r = 0.641, 95% confidence interval (CI) 0.510–0.743, p < 0.001). In a head-to-head comparison, we observed that the performance of HBcrAg was significantly superior to that of hepatitis B surface antigen (r = 0.665 vs. r = 0.475, respectively, p < 0.001). Subgroup analysis showed that the correlation between HBcrAg and intrahepatic HBV cccDNA was high, both in hepatitis B e antigen-positive and -negative patients (r = 0.678, 95% CI 0.403–0.840, p < 0.001, and r = 0.578, 95% CI 0.344–0.744, p < 0.001, respectively). In conclusion, the measurement of serum HBcrAg qualifies as a reliable non-invasive surrogate for the assessment of an intrahepatic HBV cccDNA reservoir.

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