BirT: a novel primer pair for avian environmental DNA metabarcoding
Bibliographic record
Abstract
Abstract Environmental DNA metabarcoding has become a widely used technique to detect animals from environmental samples and is on the brink of being implemented into routine species monitoring. Surprisingly, birds are among the taxonomic groups which until recently, received comparably little attention in eDNA research, a fact that is changing rapidly, with the growing number of air eDNA analyses. Since birds are hardly ever the most abundant species in aquatic or terrestrial habitats, a high specificity of the employed metabarcoding primers is key to limit non-target amplifications and enable reliable detection from environmental samples. Here, we present a novel primer pair (BirT) for metabarcoding of avian environmental DNA. We optimized specificity and fragment length regarding taxonomic resolution and available sequencing technology. Additionally, we evaluated the availability of 12S reference sequences for birds and filled database gaps by generating novel 12S barcodes. Finally, we tested the applicability of this approach using field-collected eDNA samples obtained with three different filter types. These results were compared to visual observations uploaded to eBird ( www.eBird.org ) during the sampling period. Our results confirm the suitability of the BirT primer pair for bird eDNA metabarcoding with optimized fragment length, no amplification of key non-target groups, and taxonomic resolution provided by the amplified fragment. Albeit there are still substantial gaps in the 12S reference sequence database, the analysis of bird eDNA from water samples resulted in species-level taxonomic resolution for 92% of the detected taxa. All tested filter/filtration combinations delivered similar results for total read numbers per sample (mean: 613,972 ± 340,088 SD) and species detected per sample (mean: 5.5 ± 2.3 SD). Ninety-five percent of the bird detections were highly plausible and 58% confirmed by visual observations. The majority of the detected bird species was closely associated with aquatic habitats confirming the suitability of water samples for the detection of waterfowl and species inhabiting similar ecological niches via eDNA.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.006 | 0.009 |
| Meta-epidemiology (narrow) | 0.004 | 0.003 |
| Meta-epidemiology (broad) | 0.004 | 0.002 |
| Bibliometrics | 0.004 | 0.002 |
| Science and technology studies | 0.001 | 0.001 |
| Scholarly communication | 0.001 | 0.001 |
| Open science | 0.002 | 0.001 |
| Research integrity | 0.003 | 0.003 |
| Insufficient payload (model declined to judge) | 0.007 | 0.008 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".