A fruit diet rather than invertebrate diet maintains a robust innate immunity in an omnivorous tropical songbird

1. Diet alteration may lead to nutrient limitations even in the absence of food limitation, and this may affect physiological functions, including immunity. Nutrient limitations may also affect the maintenance of body mass and key life history events that may affect immune function. Yet, variation in immune function is largely attributed to energetic trade-offs rather than specific nutrient constraints. 2. To test the effect of diet on life history traits, we tested how diet composition affects innate immune function, body mass and moult separately and in combination with each other, and then used path analyses to generate hypotheses about the mechanistic connections between immunity and body mass under different diet composition. 3. We performed a balanced parallel and crossover design experiment with omnivorous Common Bulbuls Pycnonotus barbatus in out-door aviaries in Nigeria. We fed 40 wild-caught bulbuls ad libitum on fruits or invertebrates for 24 weeks, switching half of each group between treatments after 12 weeks. We assessed innate immune indices (haptoglobin, nitric oxide and ovotransferrin concentrations, and haemagglutination and haemolysis titres), body mass and primary moult, fortnightly. We simplified immune indices into three principal components (PCs), but we explored mechanistic connections between diet, body mass and each immune index separately. 4. Fruit fed bulbuls had higher body mass, earlier moult and showed higher values for two of the three immune PCs compared to invertebrate fed bulbuls. These effects were reversed when we switched bulbuls between treatments after 12 weeks. Exploring the correlations between immune function, body mass and moult, showed that an increase in immune function was associated with a decrease in body mass and delayed moult in invertebrate fed bulbuls, while fruit fed bulbuls maintained body mass despite variation in immune function. Path analyses indicated that diet composition was most likely to affect body mass and immune indices directly and independently from each other. Only haptoglobin concentration was indirectly linked to diet composition via body mass. 5. We demonstrated a causal effect of diet composition on innate immune function, body mass and moult: bulbuls were in better condition when fed on fruits than invertebrates, confirming that innate immunity is nutrient specific. Our results are unique because they show a reversible effect of diet composition on wild adult birds whose immune systems are presumably fully developed and adapted to wild conditions – demonstrating a short-term consequence of diet alteration on life history traits.,This dataset includes information from a diet manipulation experiment on Common Bulbuls in out-door aviaries in Nigeria. We caught 40 adult Common Bulbuls using mist nets around the A. P. Leventis Ornithological Research Institute (APLORI) in Nigeria (09°52’N, 08°58’E) between 28 October to 7 November 2016 and housed them in groups of 10 birds in four adjacent out-door aviaries at APLORI. Birds were fed fruits and invertebrates in captivity until the experiment started on the 2 December 2016. Birds were supplied water and food ad libitum before and throughout the experiment. All birds were sampled for blood, assessed for moult and weighed to determine baseline body mass and innate immune function on 1 or 2 December, before diet restriction commenced on 2 December. During the experiment, birds in two aviaries were fed fruits , and the other two were fed invertebrates and sampled fortnightly. After 12 weeks of diet treatment, five birds from each aviary were switched between treatments, and the other five birds of each aviary remained on the same treatment. Switched birds replaced each other in aviaries with the alternative diet treatment, so we maintained four aviaries with the same diet treatment throughout the experiment. In one of the fruit treatment aviaries, we moved only four birds to the invertebrate treatment because we had nine birds left in this aviary. The experiment continued for another 12 weeks. Thus, we grouped individuals as: invertebrate throughout, invertebrate to fruit, fruit to invertebrate and fruit throughout. There were six females and 14 males on fruit diet and nine females and 11 males on invertebrate diet at the start of the experiment, but we were blind to the sex of individuals during the experiment, because sexes were only determined molecularly after the experiment. All birds were sexed using gel electrophoresis. Birds were sampled between 6:00 and 10:00 hours daily in two consecutive days per sampling session. Two aviaries of alternate diet treatments were sampled per day, with sampling order rotating between sampling sessions.Plasma and blood cells were stored at -20° C for one week and then moved to -80° C until transported for immune assays in Groningen, the Netherlands. Haptoglobin, nitric oxide and ovotransferrin concentration were carried by colorimetric assays, absorbance were measured using a Versamax plate reader (Molecular Devices Sunnyvale, California, US). Natural antibody-mediated haemagglutination and complement-mediated haemolysis titres of plasma samples against 1% rabbit red blood cells (Envigo RMS (UK) Ltd.) in phosphate buffered saline were measured as described by Matson et al. (2005). Additonal morphometric measurements, including wing length, tarsus length and body mass are included in the dataset. Moult status and scores of feathers for each individual are also included in the dataset. Haematocrit measurements (PCV) and occurrence of ectoparasite, and microfilaria are also recorded in the data where available, although these were not analysed for the current manuscript.,Notes are provided as comments where necessary for each column of data in the separate excel worksheets. This provides information about measurement units and scoring. Each worksheet contains separate data for either sampling information or immune assays and can be linked by the Ring number of each bird.