Great tit predation on colour morphs of the wood tiger moth at different relative frequencies

Freeze-killed adult wood tiger (Arctia plantaginis) moths from a multi-year laboratory stock, were thawed and spread so that their hindwing color was visible. The forewing length of each moth was measured with calipers prior to the start of the experiment. Specimens were then laid unpinned on their ventral side, with the dorsal side visible on individual Petri dishes. Each experiment contained a total of 24 moths laid out in a 4x6 grid in one of two enclosed indoor aviaries. The floor of each aviary was covered in dark green sheeting (tarp) to approximately mimic a natural background. Four different frequency treatments were created as follows: Control – 8 white males, 8 yellow males, 8 orange/red females (hereby just called red); Red biased- 6 white males, 6 yellow males, 12 red females; White biased – 12 white males, 6 yellow males, 6 red females; and Yellow biased – 6 white males, 12 yellow males, 6 red females. Each bird was assigned to a single treatment. Moth position on the grid was randomized. Wild Great tits (Parus major) were caught from baited traps at Konnevesi Research Station (Central Finland), where this experiment took place, in October 2015. Once trapped, all birds were measured, aged, sexed, housed individually in plywood cages (80x65x50 cm) with a daily light period of 11h:13h (light:dark), fed on sunflower seeds, peanuts and vitamin-enriched tallow, and provided with fresh water ad libitum. After the experiment, all birds were ringed for identification purposes before being released at the capture site. Wild birds were used with permission from the Central Finland Centre for Economic Development, Transport and Environment, licensed from the National Animal Experiment Board (ESAVI/9114/04.10.07/2014) and the Central Finland Regional Environment Centre (VARELY/294/2015), and used according to the ASAB guidelines for the treatment of animals in behavioral research and teaching. Birds were trained in groups overnight to forage in the experimental room and take palatable food (peanuts and sunflower seeds) from the petri dishes laid out in the grid, before being returned to their home cages. Trials were run the following day after training. Prior to the start of each trial the participating bird was food deprived for one to two hours to ensure they were motivated to forage. Twenty-four moths were then laid out on petri dishes in a 4x6 experimental grid (Figure 1) and a single bird was released. We observed each trial through a one-way mirror and recorded the timing, order, and outcome (i.e. eaten or rejected/dropped) of each attack, as well as other bird behaviors such as beak wiping or cleaning. Trials lasted until each bird had attacked at least twelve moths, or two hours had passed, whichever came first. All treatments included 10 trials and birds were only used for a single trial, for a total of 40 birds used in the entire experiment. Following their use in the experiment, birds were given at least six hours to feed in their home cages before being released.