Molt and the Immune System of Laying Hens

Feathers are complex structures essential to a bird’s survival which must be periodically replaced by molting. The process of molt in birds is characterized by multiple physiological changes and is known to be expensive in terms of the nutrients required to produce quality feathers quickly. However, little is understood about the relationship between the costly processes of molt and immune function. The overall objective of this dissertation was to determine how molt affects the immune system in regard to both systemic shifts and a potential trade-off between molt and mounting an innate immune response. In the first experiment, we investigated how the spleen, thymus, and inflammatory state of the domestic chicken (Gallus domesticus) shifts over the course of molt. Half of the adult laying hens (N = 36) were induced to molt using a combination of oral thyroxine (T4), a reduced calorie diet and a shift from a long day to a short-day schedule. Blood, liver, spleen, and thymus samples were collected weekly to assess changes over the course of molt. To determine if molt induced either a pro- or anti-inflammatory state, the level of key pro- and anti-inflammatory cytokines were evaluated via qPCR and ELISA. The spleens of molting birds were 63% heavier than the spleens of non-molting birds, and the thymus weight of molting birds increased 6.8-fold (P < 0.01). Upon histological examination, the thymus of molting birds had clearly defined lobules and an increase in cortex and medulla tissue (P < 0.01). Molting birds had a significantly higher proportion of double-positive T cells in the thymus than non-molting birds (P < 0.01). Overall, the increase in mass of both the thymus and the spleen resulted in molting birds having a greater number of T cells. Both IL-8 and TNF-alpha expression increased in the liver during molt, but overall, there was no clear indication that molt leads to an increase in inflammation during feather growth. During molt, there are marked changes in the weight and lymphocyte populations of the spleen and thymus which may be driven by molt or triggered by the hormonal shifts that occur with molt. In the second experiment, molting birds were subjected to an innate immune challenge to clarify how the nutritional requirements of molt may alter investment in an immune response. Adult hens (N = 16) were induced to molt using the same combination of cues described for experiment one. During week four of molt, birds were challenged with an intra-abdominal injection of lipopolysaccharide (1.5 mg/kg) to induce an innate immune response. Four hours post injection, liver, spleen, and blood samples were collected to evaluate changes in the expression levels and plasma concentration of key pro- and anti-inflammatory cytokines. The weight of the spleen and thymus were measured to assess changes in lymphoid tissue mass during molt. The spleen and thymus were both significantly heavier in molting birds than non-molting birds (P < 0.01). Based on inflammatory cytokine expression levels in the liver and spleen, both molting and non-molting birds mounted an innate immune response to LPS, but the response was dampened in molting birds. The expression levels of IL-6 and IL-8 were significantly reduced in molting birds. The concentration of IL-1-Beta and IL-6 in the plasma of molting birds was also significantly reduced compared non-molting birds indicating that production of protein as part of the inflammatory response may be attenuated due to competing demands for protein deposition in feathers. The third experiment used a unique genetic line, the Scaleless High (ScHi) line, to further examine the trade-off between molt and an innate immune response by mitigating the cost of feather production. ScHi birds have reduced feathering due to a genetic mutation and were used to determine if the trade-off between molt and the immune system is driven mainly by the mass of feathers grown during molt or by the broader process of molt and the accompanying hormonal and metabolic changes. Adult hens from both genetic lines (N = 32) were induced to molt using the combination of hormonal, light, and dietary signals. During week four of molt, half of the birds in each genetic line were challenged with an intra-abdominal injection of lipopolysaccharide (1.5 mg/kg) to induce an innate immune response. Five hours after the LPS injection, samples were collected from the liver and spleen to evaluate the expression of pro- and anti-inflammatory cytokines. The weights of the spleen and thymus were measured to determine if systemic changes in these lymphoid tissues during molt were dependent on the mass of feathers growing during molt. The spleen and thymus significantly increased in weight during molt in both genetic lines (P < 0.01). The thymus weight increased 7.8-fold while the spleen weight increased an average of 48% across molting birds from both genetic lines. The area of cortex in the thymus unexpectedly increased more in molting reduced feathering ScHi birds than in the fully feathered UCD-003 birds. The inflammatory response to LPS was dampened in both genetic lines, especially the expression levels of IL-6 and IL-8. The overall process of molt, rather than the mass of feathers grown, appears to drive the trade-off between molt and the immune system when both processes must occur concurrently. This research helps clarify how a nutritionally expensive component of immunity, the inflammatory response, is affected by molt when birds must balance the costs of the immune system with the costs of molting. When molt and an innate immune response must occur concurrently, the inflammatory response is dampened, likely as a mechanism to allow both molt and an innate immune response to progress during an energetic or nutritional bottleneck