Animal models

Seventy non- or post-lactating adult female greater mouse-eared bats (M. myotis) were caught at their home roost (Orlova Chuka Cave, district Ruse) in Northeast Bulgaria, between 12 July and 7 August, 16–22 h before the experimental treatment and were kept at the nearby Siemers Bat Research Station, Tabachka. All experiments were carried out under the licence of the responsible Bulgarian authorities (the Bulgarian Ministry of Environment and Water, and Regional Inspectorate (RIOSV) Ruse, permit #465/29.06.2012).

Experimental treatment

For the experiments, bats were put in holding cages at a treatment site 1.3 km away from their cave and offered a clear view of the horizon in all directions from 22 min before until 75 min after sunset (when the last visible post-sunset glow had disappeared). All experimental evenings generally had clear sky and always had a visible sunset with only the exceptional cloud coming up. Bats were placed in holding cages which consisted of an inner (120 × 120 × 63 mm) and outer (148 × 148 × 72 mm) cardboard box. The inner holding box had windows (98 × 47 mm) covered with plastic mesh (4 mm square). The outer experimental box also had windows (108 × 52 mm), covered with an outer layer of a pseudo-depolarizing filter (90% depolarization with a 10–15% reduction of light intensity for a range of 400–800 nm)6, effectively eliminating natural polarization. The windows further had an inner layer which was a polarizing filter (linear polarizer P500, 3Dlens Corporation Taiwan, ultraviolet block, transmittance: 43%, polarizing: efficiency 99.9% at 380–700 nm)2. The polarization direction of the inner filter was either vertically or horizontally oriented, with opposite sides having the same direction (Fig. 2). The boxes were oriented either with the vertically polarized windows 90° away from the sun (in a North–South axis), corresponding to the natural situation (PN), or they were shifted 90° so that now the horizontally polarized windows were oriented North–South (PS) (Fig. 1a). Bats were also kept in a double-wrapped magnetic Helmholtz coil with current antiparallel (resulting in no change of the natural magnetic field), as they served as a control group for another experiment (see Supplementary Data 1 for details of release nights).

Figure 2: Experimental boxes. On the left is the outer box (polarization box) with two different layers of filters. On the outside is a pseudo-depolarizing filter and on the inside a polarizing filter. The direction of polarization is indicated with arrows. On the right is the inner box (holding box) with its meshed windows. Full size image

Testing

After the treatment, the bats were translocated to either of two release sites, with RS1 23.6 km north–north–east and RS2 20.4 km south–west–west away from the treatment site. Both sites were flat fields, offering a clear line of sight in all directions. Bats were then fed 20–40 mealworm larvae (Tenebrio molitor) and watered ad libitum. They were equipped with radio transmitters (BioTrack PicoPip AG379, 0.44 g) and released singly starting at 0100 hours. The bats were tracked from the roof of a car using a radio receiver (AR8200 III, AOR) connected to a five-element Yagi antenna which was mounted on a 4 m pole, thus reaching ~6 m in height. The release direction of the bats was chosen randomly and the only person tracking them was blind to the treatment. Each bat was tracked until the radio signal could no longer be heard and after at least 1 min of silence the direction was noted as the vanishing bearing. Each evening the same number of bats for PN and PS were released with 15 each for RS1 and 16 PN and 17 PS for RS2. For RS1 we added another control group which received no polarization treatment (CC, n=14).

Statistical analysis