# Applicability of Non-Invasively Collected Eurasian Goshawk (Astur gentilis) Moulted Feathers for Whole Genome Sequencing Analysis

## TL;DR

The study shows that moulted feathers from Eurasian goshawks can be used for whole-genome sequencing, though sample quality varies and careful selection is needed.

## Contribution

Demonstrates the feasibility of using non-invasive moulted feathers for genome sequencing in raptors, highlighting factors affecting DNA quality.

## Key findings

- Feathers with blood traces and larger size yielded better DNA quality and sequencing performance.
- Approximately 83% of the genome was covered at least once, with millions of genetic variants identified.
- About 22.7% of samples failed due to poor DNA quality or high missing data.

## Abstract

Background/Objectives: Non-invasive samples offer an attractive alternative to logistically challenging invasive approaches in wildlife genetic studies but often contain low-quality host DNA that limits downstream analyses. Here, we assessed the applicability of moulted Eurasian goshawk feathers as a DNA source for whole-genome re-sequencing. Methods: We analysed 75 moulted feathers collected opportunistically from breeding territories. Each feather was measured from tip to tip, and its condition was visually assessed. Whole-genome re-sequencing was performed with a target coverage of 13× using 150 bp paired-end reads. Results: Feathers yielded an average of 7.19 ± 10.93 ng/μL DNA. DNA yield was positively correlated with feather size and the presence of blood traces in the calamus. On average, feather samples performed well, producing 208.7 ± 59.82 million reads, of which 82.69 ± 27.15% aligned to the reference genome, resulting in 83.58 ± 19.02% of the genome being covered at least once. After quality filtering, 10.34 ± 3.11 million biallelic single-nucleotide variants remained, of which 457,745 were common variants (MAF > 0.05). Larger feathers in good condition, with higher DNA yields and blood traces in the calamus, tended to perform better throughout the re-sequencing workflow. Nevertheless, approximately 22.7% of samples failed due to high missing data or poor genotype quality. Conclusions: Performance varied substantially even among samples with similar characteristics, indicating that improved sample selection incorporating direct measures of host DNA quality may be beneficial. Despite these challenges, moulted feathers represent a readily available DNA source for genome-wide re-sequencing of medium- to large-sized raptor species.