Bead solution

The preliminary step of the DNA extraction process involved the use of bead beating with 0.1 mm diameter glass beads (BioSpec Products, Bartlesville, OK USA) on a Powerlyser 24 homogenizer (Mo-Bio, Carlsbad, CA USA) at the Australian Centre for Ecogenomics (ACE), Brisbane, Australia. Briefly, samples were transferred to a bead tube and 800 µl of bead solution (Qiagen, Germantown, MD USA) was added and bead-beat for five minutes at 2000 rpm, then centrifuged at 10,000 g for one minute. Following the addition of 60 µl of cell lysis buffer, tubes were vortexed and then heated at 65 °C for 10 min (while mixing at 1000 rpm), then vortexed again for 30 s and stored at − 20 °C pending DNA extraction. Prior to DNA extraction, samples were thawed at room temperature; vortexed and centrifuged for one minute at 10,000 g. The resulting lysate was transferred to a new collection tube and DNA extraction carried out using DNeasy Powersoil Kit (Qiagen), as per manufacturer protocol with a final elution volume of 50 µl using sterile, EDTA-free elution buffer.

The preliminary step of the DNA extraction process involved the use of bead beating with 0.1 mm diameter glass beads (BioSpec Products, Bartlesville, OK USA) on a Powerlyser 24 homogenizer (Mo-Bio, Carlsbad, CA USA) at the Australian Centre for Ecogenomics (ACE), Brisbane, Australia. Briefly, samples were transferred to a bead tube and 800 µl of bead solution (Qiagen, Germantown, MD USA) was added and bead-beat for five minutes at 2000 rpm, then centrifuged at 10,000 g for one minute. Following the addition of 60 µl of cell lysis buffer, tubes were vortexed and then heated at 65 °C for 10 min (while mixing at 1000 rpm), then vortexed again for 30 s and stored at −20 °C pending DNA extraction. Prior to DNA extraction, samples were thawed at room temperature; vortexed and centrifuged for one minute at 10,000 g. The resulting lysate was transferred to a new collection tube and DNA extraction carried out using DNeasy Powersoil Kit (Qiagen), as per manufacturer protocol with a final elution volume of 50 µl using sterile, EDTA-free elution buffer.

Foals were sampled on a once weekly schedule from birth. Rectal swab samples were taken from foals once a week until the foal was either 5 or 6 weeks old. All ten SFM foals were sampled weekly through week 6. Six DCM foals were sampled through week 6 and the remaining 4 foals were sampled through week 5 due to the inability to access them for sampling during their sixth week of life. Each dam was sampled once at week 5 or 6 post-partum during the study period. Swab samples were collected in triplicate using sterile cotton-tipped swabs, stored on ice for no more than an hour, then placed in a bead tube containing 750 μl of bead solution (MO BIO Laboratories Inc., Carlsbad, CA). The tubes were then stored in a freezer at -20 °C until extraction.