Summary
Individual variation in parasite defences, such as resistance and tolerance, can underlie heterogeneity in fitness and could influence disease transmission dynamics. Glucocorticoid hormone concentrations often change in response to fluctuating environmental conditions and mediate changes in immune function, resource allocation and tissue repair. Thus, changes in glucocorticoid hormone concentrations might mediate individual variation in investment in resistance versus tolerance. In this study, we experimentally increased glucocorticoid concentrations in red-winged blackbirds (Agelaius phoeniceus) that were naturally infected with haemosporidian parasites, and assessed changes in resistance and tolerance of infection. Glucocorticoid treatment increased burdens of Plasmodium, the parasite causing avian malaria, but only in the absence of co-infection with another Haemosporidian, Haemoproteus. Thus, glucocorticoids might reduce resistance to infection, but co-infection can mitigate the negative consequences of increased hormone concentrations. Glucocorticoid treatment also decreased tolerance of infection. We found no evidence that the inflammatory immune response or rate of red blood cell production underlie the effects of glucocorticoids on resistance and tolerance. Our findings suggest that exogenous glucocorticoids can increase the costs of haemosporidian infections by both increasing parasite numbers and reducing an individual's ability to cope with infection. These effects could scale up to impact populations of both host and parasite.
Methodology
Study population
Red-winged blackbirds breeding in the marshes at the Queen's University Biological Station (QUBS; 44°34′02.3″ N, 76°19′28.4″ W) and in the surrounding Rideau Canal region of southeastern Ontario, Canada, have high haemosporidian infection prevalence. In the late 1980s to early 1990s, blood smear analyses revealed that 30–71% of birds in the QUBS population tested positive for the parasites [26–28]. In 2013–2015, PCR-based screening of breeding adults indicated that haemosporidian prevalence was 97.97 ± 4.55% (mean ± s.e.) and 64.73 ± .43% (mean ± s.e.) were infected with parasites from at least two genera of haemosporidians [13].
Experimental design and data collection
In April–May 2015, we captured 89 adult males either using V-top Troyer traps baited with seeds and conspecific decoys or mist nets paired with conspecific song and call playback. We housed birds in groups of three in large outdoor flight aviaries (2.4 × 6.1 × 2.4 m, 30 aviaries in the complex; see the electronic supplementary material, Methods and Results, for detailed information about bird capture and husbandry).
We randomly assigned birds to one of three treatment groups, such that each aviary contained one bird from each treatment group, with the exception of one aviary containing only two birds. Treatment groups were low-dose corticosterone (0.1 mg implant, n = 30), high-dose corticosterone (0.5 mg implant, n = 30) and control (vehicle-only implant, n = 29) (Innovative Research of America, Sarasota, FL, USA). We sampled birds once pre-treatment, then immediately implanted the pellets subcutaneously on the birds' backs. We sampled each bird again 7, 14 and 21 days post-treatment. Corticosterone doses and sampling periods were selected based on a pilot study conducted in 2014 (methods and results for the pilot study are located in the electronic supplementary material). At each sampling period we collected 500 µl of blood and measured body mass. We used blood samples to assess each individual's health, immune function, and tissue repair, and parasite presence and burdens. We evaluated host health using four metrics that can be impaired by haemosporidian infection: haematocrit [22], haemoglobin [22], body mass [29] and oxidative balance [30,31]. We also assessed an indicator of genomic stability, the percentage of red blood cells containing micronuclei [32,33]. Micronuclei are formed when chromosomes break or are not incorporated into the nucleus during cell division [32]. Birds infected with haemosporidian parasites upregulate red blood cell production to compensate for the cells destroyed during infection [22], and this increase in cell production might result in higher error rates during cell division. We also assessed two immune metrics that can increase in response to haemosporidian infection: (i) nitric oxide, an inflammatory signalling molecule and anti-parasitic defence [34–36]; and (ii) PIT54 (avian analogue of haptoglobin), an acute-phase, anti-inflammatory protein and haemoglobin scavenger [37–39]. Because both haemosporidian parasites and the immune response to malarial parasites damage red blood cells [40], we assessed the level of red blood cell production as an indicator of tissue repair rate. Parasite presence/absence and burden were assessed pre-treatment and 7 days post-treatment, and all other measures were assessed at every sampling point.