Study site and sampling
The study was conducted from 2012 to 2017 in a free-living population of
jackdaws (Corvus monedul a) breeding in nest boxes located south
of Groningen, The Netherlands (53° 14’N, 6°64’E), which has been
routinely monitored since 2005. Nestboxes were routinely checked to
assess egg laying and, after incubation commenced, both pair-members
were identified by their unique colour ring combination. Brood size
manipulations are described in detail in . In brief, broods were
manipulated when the oldest nestling was 4 days old (hatching date = day
0, hereafter, day 5). Dyads of broods that were manipulated were matched
by clutch size and laying date (±1 day). Nestlings were cross fostered
so that the net manipulation was ±2 nestlings, creating enlarged and
reduced broods with both manipulated broods containing nestlings hatched
in both broods in the dyad . Biometric measurements and weight (to the
nearest 0.1 g) were taken on days 5 and 30, and a blood sample was taken
from the brachial vein. Upon collection, all blood samples were stored
in 2% EDTA buffer at 4– 7°C and within 3 weeks they underwent snap
freezing in a 40% glycerol buffer for long-term storage at −80°C.
Parasite counts (hereafter Carnus counts) were recorded for each
nestling on every visit to the nest (due to regular nest checks and the
experimental manipulation, data on parasite infestation were available
for days 0, 1, 5, 10, 11, 20 and 30). Parasites were located by visual
inspection of the nestlings’ body, with particular attention to areas
where either the parasite or bite marks are common (under the wings and
legs and belly). From 2012 to 2014 both parasite counts and bite marks
on the belly were recorded (n = 229), while from 2015 to 2017 only
parasite presence was recorded. Bite marks were recorded as a ‘belly
score’ index from 0-4 (where 0 = no marks, 1 = under 10 marks, 2 = 10-20
marks, 3 = 20-50 marks, 4 = 50 or more marks).
We validated the belly score index as indicator of parasite presence in
the nest (for cases when no Carnus flies were seen but belly
scores was > 0) using a subset of individuals for which
both parasite counts and belly score data were available. Belly score
and Carnus counts predict the same outcome (belly score = 0Carnus counts = 0 vs. belly score > 0 Carnuscounts > 0) in 94% of cases
(χ23 = 180.84, p-value <
0.0001). We further validated that belly score is a good indicator of
parasite abundance (Table S1, Fig. S1) with a generalised linear mixed
model with Poisson error distribution with Carnus counts as
dependent variable and belly score index, age, their interaction, and
year as fixed effects. This model included foster nest and colony as
random effects to control for repeated measures and spatial variation.
Because parasites often move from one sibling to another within the nest
(, we scored the whole brood as ‘infested’ when at least one nestling
from a brood was infested at any sampling point.
We recognize that variation in parasite infestation would ideally have
been experimental in origin but in this study we relied on natural
variation. This may have biased our results when Carnus infection
varies systematically between or within colonies, or when jackdaws could
detect Carnus infection when selecting a nest site, but neither
appears to be the case. That the flies are widespread and first appear
long after (egg) incubation has led us to assume for the present study
that jackdaws have little control over whether they breed in a nest box
with or without Carnus infection.