10.5061/DRYAD.CVDNCJT3C
Jamie, Gabriel
0000-0002-2766-7687
University of Cambridge
Hamama, Silky
University of Cambridge
Moya, Collins
University of Cambridge
Kilner, Rebecca
University of Cambridge
Spottiswoode, Claire
0000-0003-3232-9559
University of Cambridge
Limits to host colonisation and speciation in a radiation of parasitic finches
Dryad
dataset
2020
FOS: Biological sciences
Leverhulme Trust
https://ror.org/012mzw131
RPG-2013-251
Royal Society Dorothy Hodgkin Fellowship *
Royal Society Wolfson Merit Award*
Royal Society Dorothy Hodgkin Fellowship
Royal Society Wolfson Merit Award
2020-12-28T00:00:00Z
2020-12-28T00:00:00Z
en
193095 bytes
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CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Parasite lineages vary widely in species richness. In some clades,
speciation is linked to the colonisation of new hosts. This is the case in
the indigobirds and whydahs (Vidua), brood-parasitic finches whose
nestlings mimic the phenotypes of their specific hosts. To understand the
factors limiting host colonisation, and therefore speciation, we simulated
the colonisation of a host using cross-fostering experiments in the field.
Despite DNA barcoding suggesting that host species feed their chicks
similar diets, nestling Vidua had low survival in their new host
environment. Nestling Vidua did not alter their begging calls plastically
to match those of the new hosts, and were fed less compared to both host
chicks and to Vidua chicks in their natural host nests. This suggests that
a key hurdle in colonising new hosts is obtaining the right amount rather
than the right type of food from host parents. This highlights the
importance of mimetic nestling phenotypes in soliciting feeding from
foster parents and may explain why successful colonisations tend to be of
hosts closely-related to the ancestral one. That non-mimetic chicks are
fed less but not actively rejected by host parents suggests how selection
from hosts can be sufficiently intense to cause parasite adaptation, yet
sufficiently relaxed that parasitic chicks can sometimes survive in and
colonise new host environments even if they lack accurate mimetic
phenotypes. The difficulties of soliciting sufficient food from novel
foster parents, together with habitat filters, likely limit the
colonisation of new hosts, and therefore speciation, in this parasite
radiation.
Transfer experiments simulating host colonisation During January–April
2014–2017, we carried out transfer experiments within an area of about 40
km2 on and around Musumanene and Semahwa Farms (centred on 16°47′S,
26°54′E) in the Choma District of Zambia. The habitat is a mixture of
miombo woodland, agricultural fields and seasonally-flooded grasslands.
The experiment had three treatments: (i) pin-tailed whydah eggs
transferred from common waxbill to blue waxbill nests, (ii) blue waxbill
eggs transferred to blue waxbill nests, and (iii) common waxbill eggs
transferred to blue waxbill nests. Additionally, we monitored survival and
feeding of pin-tailed whydah nestlings in naturally-parasitised common
waxbill nests. To minimise predation, eggs were taken from their natural
nest and incubated in a Brinsea Octagon 20 Advance EX Incubator at 36.7°C
and 60% humidity. A day before they were due to hatch, eggs were fostered
to a blue waxbill nest. Occasionally (16/94 transfers), the offspring had
to be fostered as a chick freshly hatched in the incubator, rather than as
an egg but this was not found to influence the offspring’s subsequent
survival in the novel nest environment (see Results). The modal clutch
size of both common waxbill and blue waxbill nests was five. No host egg
was removed when an egg/hatchling was added, because pin-tailed whydah
females do not remove a host egg when they naturally parasitise a
nest (Tarboton, 2011). Experimental nests were visited every two or three
days, and the number of eggs and chicks in the nest was recorded. For
chicks, mass and tarsus length were measured and the amount of food in the
crop scored (methods below). Mass was measured on digital scales to an
accuracy of 0.1 or 0.01 g depending on the model of scale. Tarsus length
was measured using dial callipers to the nearest 0.1 mm. All three
species used in the study are common and widespread and experience high
levels of natural nest predation such that the experiments carried out for
this study will have negligible effects on their populations. Data were
collected under the research approval of the Department of National Parks
and Wildlife in Zambia (DNPW/8/27/1). Our sample sizes were chosen to
allow the between-species effects to be detected with a high degree of
confidence while not including an unnecessarily high number of
individuals. As such it meets the ABS/ASAB guidelines and adheres to the
three R’s of replacement, reduction and refinement (Buchanan et al.,
2012). Comparing survival of different species transferred to blue waxbill
nests Survival analyses were carried out in the R statistical
environment(R Development Core Team, 2017) using the packages
Survival (Therneau, 2015) and KMsurv (Moeschberger and Yan, 2012). We
monitored chick survival from the day the chick hatched in the new host
nest. Chick survival was judged to end at the midpoint between the last
day the chick was known to be alive and the first day the chick was known
to be absent. If the nest was still active, but the transferred chick was
absent, then the chick was assumed to have died. If the nest was abandoned
at the point the transferred chick was absent, then the data were
right-censored. Right-censoring is used when the event of interest has not
occurred by the last observation (Mills, 2011). A Cox proportional hazards
model was fitted to the survival data (Cox, 1972). The co-variates in the
initial model were: (i) transferred chick species, (ii) presence of host
nestmates, and (iii) foreign chick transferred as egg or as chick. The
number of nestmates over the course of a given transfer experiment ranged
from 0 to 5 (mean = 1.4). In most transfers (49 of 94), the transferred
chick had no nestmates in the foster nest. We therefore modelled nestmate
presence as presence/absence. Comparing the amount of food host parents
fed transferred chicks of different species Crop size of the transferred
chick was recorded at each nest visit. Nestling estrildid crops are
transparent, allowing easy visual inspection. Crops were scored as 0
(empty), 1 (trace amounts, < c. 20 seeds, no bulge in crop), 2
(> c. 20 seeds, slight bulge) or 3 (> c. 50 seeds, large
bulge). To assess whether crop sizes of chicks differed depending on the
species of chick transferred, two approaches were used: First, the
median crop size of the transferred chick over the first 7 days of
survival in the host nest was used as the response variable; c. 80% of
common waxbill and pin-tailed whydah chick mortality occurred in this
period (Figure 2). A Kruskal-Wallis test was used to test whether median
crop size differed between the three species. Median crop sizes were
compared between species using a Dunn post-hoc test, using the dunn.test
function from the R package dunn.test and with Bonferroni correction for
multiple testing (Dinno, 2017). Second, ordinal mixed-effect models
were implemented in the R package Ordinal (Christensen, 2015) using crop
score as an ordinal response variable. In the full model, the fixed
effects were chick species and the number of nestmates. Transferred chick
individual was a random effect nested within the nest of origin of that
transferred chick. We carried out stepwise elimination of non-significant
co-variates until only significant co-variates remained. The model was
initially run to include crop scores over the first 7 days of development,
then re-run using crop scores over the first 4, 5, 6 and 8 days of
development to test whether the findings from the first 7 days of
development were robust. Ordinal mixed-effect models were used to
compare crop size in pin-tailed whydahs and common waxbills occurring in
their natural nests (common waxbill nests) with those experimentally
transferred to blue waxbill nests. Data for pin-tailed whydahs and common
waxbills in common waxbill nests are observational, unlike the
experimental data from pin-tailed whydahs in blue waxbill nests. This was
because high levels of nest predation meant that all common waxbill nests
found at the egg stage were required as a source of eggs for transfer to
blue waxbill nests. Therefore, our data on pin-tailed whydahs in common
waxbill nests do not account for the effects of transferring an egg from
one nest to another. However, when blue waxbill eggs were transferred from
their own nest to another blue waxbill nest, they still showed high
survival (about 76% of all chicks transferred survived to fledging),
suggesting that any effects of transferring eggs between nests are
insufficient to account for the differences observed in chick survival
between common and blue waxbill nests. Measuring nestling diet
composition Obtaining crop samples in field Nestling crops were sampled
using the tube insertion method (Zann and Straw, 1983). The tube was
inserted in the throat of the nestling and seeds were pushed from the
translucent crop into the tube. The contents were stored in 70% ethanol.
The process was repeated until about 20–30 seeds had been extracted.
Chicks were sampled around the time when the primaries first erupt from
pin (approximately 6–7 days of age). We sampled crops of common waxbill,
blue waxbill as well crops of six other sympatric estrildid finch species:
orange-winged pytilia (Pytilia afra), melba finch (Pytilia melba),
Jameson’s firefinch (Lagonosticta rhodopareia), red-billed firefinch (L.
senegala), African quailfinch (Ortygospiza atricollis) and bronze mannikin
(Spermestes cucullatus). We sampled these other estrildid finch species in
addition to the two used in the experiments to assess variation in host
diet across a broader phylogenetic scale and to explore whether estrildids
that hosts to Vidua have a different diet from those that are not. DNA
barcoding of nestling crop contents Nestling estrildid finch crops
contained almost exclusively plant seeds. DNA barcoding of samples was
carried out by Jonah Ventures (Boulder, Colorado; jonahventures.com). The
chloroplast trnL intron was amplified from DNA in each sample using the c
and h trnL primers (Taberlet et al., 2007). The total expected amplicon
length was 332bp (Jonah Ventures in litt.). A detailed protocol is
described in supplementary methods. We consulted with an expert botanist
based in Zambia, Mike Bingham, to validate the taxonomic identities
assigned by DNA barcoding. Quantifying crop contents DNA barcoding data
resolution allowed analysis at the subfamily level and not the genus
level, so each OTU was assigned to one of the four subfamilies identified
(see Results). For each sample, reads from OTUs mapping to the same
sub-family were summed together to give a measure of the total number of
reads from each subfamily, and expressed as a proportion of total
reads (Craine et al., 2015; Willerslev et al., 2014). To test whether
different estrildid species fed chicks different proportions of seeds from
each of the four families, non-metric dimensional scaling (NMDS) was
performed using the R package vegan (Oksanen et al., 2017). Comparisons of
diet between species were made using the function adonis, from the R
package vegan. Begging call plasticity Recording nestling begging calls
Chicks were placed in an artificial nest, and given several minutes for
acclimation. To stimulate begging, the chick was tapped gently with
forceps on the bill. Recordings were made using an Audio-Technica ATR35S
tie-clip microphone or a Sennheiser ME-66 shotgun microphone held
approximately three cm away from the chick’s mouth. Recordings were made
for around two minutes or until at least ten seconds of continuous begging
were recorded. Analysing the effect of host environment on nestling
begging calls We compared begging calls of nestling pin-tailed whydahs in
natural common waxbill nests, to those transferred to blue waxbill nests.
We identified four distinct call types both by listening to recordings and
through visual inspection of sonograms (see Results). All four call types
were detected in both pin-tailed whydahs developing in common waxbill
nests, and pin-tailed whydahs transferred to blue waxbill nests. To
analyse whether host environment influenced the stage in the nestling
period at which each call type was made, we examined the stage in
development at which chicks made each call type and compared this between
pin-tailed whydahs raised in common and blue waxbill nests. We used chick
tarsus length as a proxy for developmental stage, because for pin-tailed
whydahs in their natural nests, the exact age in days of the chick was
unknown, whereas tarsus length was available for all treatments. We
examined whether within each call type, there were changes in call
structure between host environments. For each call type, the following
begging call parameters were extracted from each recording: minimum
frequency, maximum frequency, centre frequency, peak frequency, frequency
bandwidth, call duration, average entropy, and energy. These are widely
used in the literature to characterise begging sounds (Anderson et al.,
2009; Butchart et al., 2003; Langmore et al., 2008). For each recording,
ten sequential call notes in a bout of begging were selected and call
parameters extracted. Call notes were not selected if they overlapped with
interfering background noises, or if they were incomplete calls. The
relationship between the call types was visualised using linear
discriminant function analysis with the R package MASS (Venables and
Ripley, 2002). We calculated call rate by dividing the dividing the number
of calls in the bout by the duration of the bout. Two approaches were
used to compare the structure of each call type between pin-tailed whydahs
raised in common waxbill nests, and those raised in blue waxbill nests.
First, a series of linear mixed models were constructed, with each call
parameter as a separate response variable. Host environment and crop size
were fitted as fixed effects and individual chick identity as a random
effect. Crop size was used as a proxy for chick hunger. When estrildid
finch nestlings are fed, they store their seed in the crop before passing
it on to the stomach. By measuring the amount of food stored in the crop,
we could assess how much the chick had recently been fed in a non-invasive
manner. Crops were scored as 0 (empty), 1 (trace amounts, < c. 20
seeds, and with no bulge in crop), 2 (> c. 20 seeds and with slight
bulge) or 3 (> c. 50 seeds and with large bulge). We controlled for
multiple testing using Bonferroni correction (Dunn, 1961). Second, we
carried out a logistic regression analysis using the R package
nnet (Venables and Ripley, 2002), allowing all call parameters to be
considered at once.