General

We studied native Turdus thrush populations in Olomouc, Czech Republic (49°35'8″N, 17°15'3″E) in 2009–2014 and introduced thrush populations in Auckland, New Zealand (36°50′26″S, 174°44′24″E) in 200914,24. Blackbirds and song thrush show very similar nest placement and habitat selection, including our study sites in both Czech Republic25,26 and New Zealand (own unpublished data from several studies12,14,27,28,29). Specifically, in all populations we sampled nests in similar habitats, from conifers and dense shrubs situated in public parks and gardens where nests of both species were interspersed. Turdus thrushes in Europe only rarely raise common cuckoo (Cuculus canorus) chicks successfully12,30,31. Current evidence suggests that blackbirds are conspecific parasites14.

Despite extensive and long-term mist-netting and colour-banding effort32, the exact identity of each tested female (only females eject in our study population27) was not always known. Therefore, we avoided sampling the same location after experimentation to reduce the chance of testing the same individuals more than once. However, prior experience with experimentation would not necessarily cause females to be more or less likely to flush on subsequent visits and empirical data from the same egg model type showed that prior experience had only weak and statistically non-significant effect on ejection probability27.

We conducted this research in accordance with the Association of Animal Behaviour and the Animal Behavior Society guidelines for the treatment of animals in research. In the Czech Republic our research methods and protocols were approved by the Research Ethics Committee of Palacký University (45979/2001–1020) and the research was conducted under licenses from the Department of Environment of the City of Olomouc (SmOl/ZP/55/6181 b/2009/Pr and SMOVZP/55/8542/2011/Kol). Although no specific permissions were required to study these two model invasive species in New Zealand, this research was approved by the University of Auckland's Animal Ethics Committee (AEC/09/2006/R512).

Fleeing distance

In 2014, in Olomouc, JH quantified how far female blackbirds and song thrush flew from their nests after flushing (to the nearest meter, up to 30 meters). Specifically, JH slowly walked to the nest until the female left, or, if necessary, he slowly moved a hand or mirror toward the nest. Then, JH estimated the average distance between the female and the nest between 10 and 20 s after flushing. This corresponds to when experimental parasitism events occurred (see below for details). We refer to this distance as fleeing distance. This should not to be confused with flight initiation distance33, which measures the distance between an observer and animal before the animal flees. We tested whether fleeing distance, manipulation dates within season (1 = 1st January) and clutch sizes, differed between blackbirds and song thrush using Mann-Whitney U-tests. Levene's test detected no heterogeneity of variance for these variables.

Does flushing influence host responses?

Nests were monitored for six days after introducing a single non-mimetic blue model egg and models present or missing after this period were deemed accepted or ejected, respectively14,24. The mass, dimensions and spectral reflectance14,28 of these egg models closely match the cuckoo eggs that are naturally found within common redstart Phoenicurus phoenicurus nests34. We used these non-mimetic blue models, because this is the most common egg model type used across Europe and has been used in the majority of studies on these species27,28,35. Using the same model type was necessary for making meaningful and quantitative comparisons of host behavior between species and populations14,36. Desertion was not a response to artificial parasitism14,24. For each experiment we recorded whether the female was flushed from the nest cup or not when the egg model was introduced. For both species we included only nests with final clutch sizes of 4–5 eggs in CZ and 3–4 eggs in NZ, which are typical in these populations29.

We ran separate statistical models for blackbirds and song thrush. We used Fisher Exact tests to determine if flushing related to host response to the model egg and present the associated odds ratios (OR) and confidence intervals (CI). We used generalized linear models to examine if flushing (yes or no) predicted host responses (binomial distribution) and latency to ejection (Poisson distribution). We controlled for other relevant variables14,24,29: year (categorical), the laying date of the first egg (continuous; 1 = 1st April in CZ and 1 = 1st September in NZ), geography (categorical; CZ or NZ), nest age (continuous; days) and clutch size (continuous). Laying date was centred for each year and geographic location (CZ or NZ) separately to remove potentially confounding effects of annual and seasonal variation14,24,29. We used Nagelkerke's R2 to estimate model fit37 for models with binomial responses and the difference between the null deviance and residual deviance divided by the null deviance (hereafter, pseudo R2) for the models with Poisson error distributions38. We performed model selection through backward elimination of non-significant terms. All of these analyses were conducted in R version 3.1.1.

Reanalysis of published data

To examine whether flushing could have confounded previously published analyses and conclusions, we reanalysed our own previously published results using the same statistical approach14 (i.e. excluding flushing) and also including flushing as an additional predictor for both the blackbird and the song thrush. In these reanalyses, we used only data for the same non-mimetic blue egg models, because we did not record flushing data for other egg model types.

Our primary interest in these reanalyses was in the effects of flushing (binary predictor) and geography (categorical 3-level predictor: sympatry, micro-allopatry and macro-allopatry with the common cuckoo Cuculus canorus, a rare interspecific brood parasite of Turdus thrushes in Europe but not in New Zealand, for definitions see Ref. 14) on egg rejection (binary response variable). In addition to flushing and geography (see above), all statistical models included nest stage (categorical predictor with four levels: egg laying, 1–3 days of incubation, 4–9 days of incubation, 10 days of incubation to hatching), laying date (first egg laid; continuous), clutch size (clutch size at clutch completion; continuous). Laying date was centred (see above) within each year for CZ and NZ separately to remove confounding effects of between-year variation of seasonal breeding and timing of experiments. We did not include egg model and its interaction with geography as in the previous analyses14 because only responses to blue models were included (see above). Otherwise, we controlled for all variables that were previously included in these models14.

We selected final models by backward elimination of non-significant terms, retaining two main factors of interest (geography and flushing) in the model regardless their significance, following the previously published methods12,39. For consistency with previously published work, we report test statistics and P-values for non-significant terms from backward elimination procedure just before the particular term was removed from the model. These reanalyses were conducted in SAS version 9.2.