Discussion
House sparrows from the introduced range had a stronger and more
variable epigenetic response to simulated infection within 8-hours
compared to individuals from the native range. Between 0- and 8-hours
post simulated infection, DNA methylation differed at more genomic
locations, with a greater magnitude of change, in introduced than native
house sparrows. These changes in DNA methylation occurred in both
directions, with some sites gaining methylation and others losing
methylation. Introduced individuals had more unique genomic locations (n
= 35) that were differently methylated, while only a single unique
genomic location was differently methylated in native individuals.
Introduced individuals had a higher variance in the magnitude of change
in DNA methylation in response to simulated infection compared to native
individuals. It is possible that the change in DNA methylation we
observed over the 8 hour time series might not reflect an immune
response per se, but could also reflect response to the stress of being
brought into captivity, or a combination of the two. These results
support the hypothesis of an epigenetically mediated invader phenotype
present among introduce house sparrows, which provided a mechanism for
plasticity in response to novel stressors (Sheldon et al. 2023).
Further, it supports the hypothesis that epigenetic buffering likely
plays a role in the manner of this response (O’dea et al. 2016; Lauer et
al. 2024).
Our results expand previous research on the importance of DNA
methylation in the response to infection in supporting the immune
response of house sparrows in multiple contexts. In response to a
parasite infection, DNA methylation differed between infected and
non-infected house sparrows, and among individuals sampled temporally
after infection, at the nestling stage (10-14 days old) and at the
fledged juvenile stage (26-37 days; Lundregan et al. 2022). Here, the
DNA methylation patterns of a selected marker gene, NR1D1, differed
between infected and uninfected individuals and was correlated to
recruitment. Our findings are congruent in detecting DNA methylation
changes post infection, yet over a much shorter timeframe. Further,
introduced house sparrows had higher expression of pathogen surveillance
genes and cytokine responses genes, to the simulated bacterial infection
investigated in the present study (McCain et al 2025). In introduced
house sparrows, as TLR-4 expression increased, IL-1β and IL-10
responses decreased, which was not detected in native sparrows. Our
results suggest that the observed differences in immune response in
introduced house sparrows are at least in part mediated by changes in
DNA methylation. In addition, introduced house sparrows with higher
epigenetic potential (estimated by the number of CpG sites in the
promoter of TLR-4 ) had higher resistance to infection bySalmonella enterica compared to individuals with lower epigenetic
potential (Sheldon et al. 2023). As epigenetic potential measures the
genetic potential for an individual to adopt different DNA methylation
states, our results suggest that both an individual’s inherent
capability to be methylated differently, and it actually being
methylated differently, are important factors in the response to
infection. Integrating these findings demonstrates that both epigenetic
potential, and the actual changing of DNA methylation state is important
in the response to infection. Also, these studies indicate that is
highly likely that the difference in change of DNA methylation we
detected between introduced and native individuals would ultimately
support introduction success.
Our results also provide new context for previous findings of the role
of DNA methylation in the success of introduced house sparrows, by
finding supporting results in how individuals change over time.
Differences in DNA methylation occur within and among introductions of
house sparrow (Liebl et al. 2013; Sheldon et al. 2018a) and these
differences manifest across the edge-core axis of introduction (Hanson
et al. 2020b). Further, DNA methylation differs between introduced and
native individuals, with those from more recent introductions having
greater variance in DNA methylation (Lauer et al. 2024). The present
temporal study found congruent results within individuals over time:
detecting differences in DNA methylation among introductions, and
between introduced and native individuals. The individual-level response
to simulated infection, suggests that the larger patterns detected may,
in part, be shaped by individual responses.
Our results also provide new information in the study of how DNA
methylation changes over time in birds. We document substantial changes
in DNA methylation state in response to simulated infection within 8
hours, which, to our knowledge, is the shortest time frame studied. In
aviary-controlled conditions, temporal changes in DNA methylation were
detected in great tits (Parus major ) among three time points, 21
days apart, across a breeding season. Time points targeted the
initiation of gonadal development, nest building, and initiation of egg
laying (Linder et al. 2021). Changes in DNA methylation in liver and
blood were correlated, and DNA methylation near transcription start
sites was correlated to decrease in gene expression. In captive great
tits from aviary conditions, changes in DNA methylation in blood were
detected both between temperature treatments and temporally across four
selected time points, which roughly spanned reproductive behaviors of
initiation of reproduction, through 50% of individuals laying eggs
(Viitaniemi et al. 2019). A relatively large number of small magnitude
changes in DNA methylation were detected and there was a large variation
in the change over time given a relative low level of methylation, with
a large amount of among individual variation. In wild collected
chestnut-crowed babblers (Pomatostomus ruficeps ), DNA methylation
in blood differed among individuals sampled at hatching, fledgling, and
1-year (Liebl et al. 2021). In this cooperative breeder, first year
dispersers had a greater number of loci that changed DNA methylation
state between hatchling and fledgling, and had lower DNA methylation,
compared to non-dispersers before fledgling but not as hatchlings or
adults. Together, these studies show within individual change in DNA
methylation is critically important to the response to environment, and
coordination of temporally variable behaviors.
While we demonstrate a clear individual response in DNA methylation to
simulated infection, it is important to note that DNA methylation is
active in multiple different contexts within individuals, and even
within cells (see Chen et al. 2022; Sheldon et al. 2022). Thus, not all
individuals in all introduced areas are expected to show identical
change in DNA methylation, or even that the response in DNA methylation
would be expected to be directional in general. Rather, it is likely
that maintaining, or increasing, the ability to change, is of primary
importance to introduced species. The potential for change and the
response to immediate local stressors might best be detected in variance
of DNA methylation among introduced individuals, or in targeted analysis
of the regulation of specific genes. Also, it is highly likely that
histone modification is another critically important epigenetic
mechanism to this process (Ray et al. 2024). We encourage investigations
in all three areas, to provide additional insights into the response of
individuals to stress and to the success of the house sparrows as
introduced species.