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.