2.1 Sample collection and DNA extraction
Samples were collected from captive live animals from two populations in 2020. Ten E-mink were sampled in captive settings at the Fundación para la Investigación en Etología y Biodiversidad (FIEB) breeding center in Spain [originated from Spanish free-ranging population (western population)] and fourteen E-mink from the conservation breeding center at Zoodyssée in France [originated from the captive EEP population (representing the eastern population), Table S1]. Two mink sampled in the Spanish breeding center were wild-born individuals but spent at least a year in captivity. One individual sampled in the Spanish breeding center was the result of a crossbreed between western and eastern mink. For MHC and microsatellite markers analysis, hair samples were collected using sterilized tweezers from each animal during a routine procedure. For the microbiota, fresh fecal samples were collected in the enclosure of each animal separately using sterilized tools and kept in 96% ethanol tubes at 4°C until further processing. As the E-mink’s diet in captivity varies by day, samples were collected at four occasions depending on the item fed to the animal the previous day. The diet of the E-mink from both breeding centers relied on 3 types of food: trout, mice and chicken.
DNA from hair samples were extracted using the DNeasy Blood and Tissue Kit from Qiagen using the manufacturer’s protocol. DNA extractions from the fecal samples collected were conducted in duplicates using the QiaAmp Mini Kit with Inhibitex (Qiagen, Germany) following the manufacturer’s instructions. Two blank extractions were made to control for contamination during the extraction process. A mock community sample (HM-783D, BEI resources) containing genomic DNA from 20 bacterial strains, at concentrations ranging from 0.6 to 1400 pg/μl, was also added in each library to confirm the reliability of our method.