Are thyroid hormones mediators of incubation temperature-induced phenotypes in birds?

Incubation temperature influences a suite of traits in avian offspring. However, the mechanisms underlying expression of these phenotypes are unknown. Given the importance of thyroid hormones in orchestrating developmental processes, we hypothesized that they may act as an upstream mechanism mediating the effects of temperature on hatchling phenotypic traits such as growth and thermoregulation. We found that plasma T3, but not T4 concentrations, differed among newly hatched wood ducks (Aix sponsa) from different embryonic incubation temperatures. T4 at hatching correlated with time spent hatching, and T3 correlated with hatchling body condition, tarsus length, time spent hatching and incubation period. In addition, the T3 : T4 ratio differed among incubation temperatures at hatch. Our findings are consistent with the hypothesis that incubation temperature modulates plasma thyroid hormones which in turn influences multiple aspects of duckling phenotype.

Thermal challenge severity differentially influences wound healing in wood duck (Aix sponsa) ducklings.

Environmental conditions during early development can profoundly influence an individual's phenotype. Development requires simultaneous maturation and orchestration of multiple physiological systems creating the potential for interaction among key systems and requiring substantial resources. We investigated the influence of thermoregulation on immunocompetence in Wood Duck ducklings (Aix sponsa). At both 1 and 2 days post hatch (dph) we evaluated ducklings' abilities to thermoregulate during a thermal challenge at one of four temperatures (36 [thermoneutral controls], 20, 10, or 5°C). At 3 dph, ducklings received a superficial wound, which was monitored until full recovery to quantify wound healing ability, an ecologically relevant, integrative measure of immune function. We demonstrated that duckling body temperature decreased with increasing thermal challenge severity, thermoregulatory ability increased with age, and thermoregulation had temperature-dependent effects on the immune system. Specifically, a more severe thermal challenge (5°C) resulted in decreased immune performance when compared to a mild challenge (20°C). We conclude that early thermoregulatory experiences are influential in shaping immune responses early in development. Furthermore, our results emphasize that future studies of environmental stressors need to consider multiple physiological endpoints since interaction among systems can result in competing physiological demands.

Incubation conditions are more important in determining early thermoregulatory ability than posthatch resource conditions in a precocial bird.

Recent research in birds suggests that investing in incubation is one mechanism by which parents can enhance the phenotype of their offspring. Posthatch environmental conditions can also shape an individual's phenotype, and it is thus possible for pre- and posthatch conditions to have interactive effects on an individual's phenotype. In this study, we examined the individual and interactive effects of prehatch incubation temperature and posthatch food availability on growth, food consumption, and thermoregulatory ability in wood duck (Aix sponsa) ducklings. Eggs were incubated at one of three temperatures (35.0°, 35.9°, or 37.0°C), and then ducklings were reared on an either ad lib. or time-restricted diet for 12 d after hatching. We found that food availability influenced duckling growth, with the slowest growth occurring in ducklings fed the restricted diet. Incubation temperature also interacted with food conditions to influence duckling growth: ducklings fed ad lib. from the lowest incubation temperature grew slower than ducklings fed ad lib. from the higher incubation temperatures. Most importantly, we found that the improvement in a duckling's ability to maintain body temperature in the face of a thermal challenge was influenced by embryonic incubation temperature but not feeding conditions. Ducklings from the highest incubation temperature experienced the greatest improvement in thermoregulatory performance with age. Our findings suggest that the prehatch environment is more important than posthatch resource conditions in determining some physiological functions and underscores the important role that incubation temperature plays in determining offspring phenotype in birds.

Embryonic developmental patterns and energy expenditure are affected by incubation temperature in wood ducks (Aix sponsa).

Recent research in birds has demonstrated that incubation temperature influences a suite of traits important for hatchling development and survival. We explored a possible mechanism for the effects on hatchling quality by determining whether incubation temperature influences embryonic energy expenditure of wood ducks (Aix sponsa). Because avian embryos are ectothermic, we hypothesized that eggs incubated at higher temperatures would have greater energy expenditure at any given day of incubation. However, because eggs incubated at lower temperatures take longer to hatch than embryos incubated at higher temperatures, we hypothesized that the former would expend more energy during incubation. We incubated eggs at three temperatures (35.0°, 35.9°, and 37.0°C) that fall within the range of temperatures of naturally incubated wood duck nests. We then measured the respiration of embryos every 3 d during incubation, immediately after ducks externally pipped, and immediately after hatching. As predicted, embryos incubated at the highest temperature had the highest metabolic rates on most days of incubation, and they exhibited faster rates of development. Yet, because of greater energy expended during the hatching process, embryos incubated at the lowest temperature expended 20%-37% more energy during incubation than did embryos incubated at the higher temperatures. Slower developmental rates and greater embryonic energy expenditure of embryos incubated at the lowest temperature could contribute to their poor physiological performance as ducklings compared with ducklings that hatch from eggs incubated at higher temperatures.

Slight differences in incubation temperature affect early growth and stress endocrinology of wood duck (Aix sponsa) ducklings.

Early developmental experiences, such as incubation conditions, can have important consequences for post-hatching fitness in birds. Although the effects of incubation temperature on phenotype of avian hatchlings are poorly understood, recent research suggests that subtle changes in incubation conditions can influence hatchling characteristics, including body size and condition. We designed an experiment to explore the effects of incubation temperature on hatching success, survival to 9 days post hatch, growth and the hypothalamo-pituitary-adrenal (HPA) axis in wood ducks (Aix sponsa). Wood duck eggs were collected from nest boxes and experimentally incubated at three temperatures (35.0, 35.9 and 37.0 degrees C), each falling within the range of temperatures of naturally incubated wood duck nests. Survival and growth were monitored in ducklings fed ad libitum for 9 days post hatch. In addition, baseline and stress-induced plasma corticosterone concentrations were measured in 2 and 9 day old ducklings. Hatching success and survival to 9 days was greatest in ducks incubated at the intermediate temperature. Ducklings incubated at 35.9 degrees C and 37.0 degrees C had 43% higher growth rates than ducklings incubated at 35.0 degrees C. In addition, ducklings incubated at 35.0 degrees C had higher baseline (17-50%) and stress-induced (32-84%) corticosterone concentrations than ducklings incubated at 35.9 degrees C and 37.0 degrees C at 2 and 9 days post hatch. We also found a significant negative correlation between body size and plasma corticosterone concentrations (baseline and stress-induced) in 9 day old ducklings. To our knowledge, this is the first study to demonstrate that thermal conditions experienced during embryonic development can influence the HPA axis of young birds. Our results illustrate that subtle changes (<1.0 degrees C) in the incubation environment can have important consequences for physiological traits important to fitness.

Evaluation of field and laboratory protocols used to detect avian influenza viruses in wild aquatic birds.

Careful selection and observance of standard field and laboratory protocols are critical for successful detection and characterization of avian influenza viruses (AIV) from wild birds. Cloacal swabs were collected from hunter-killed or nesting waterfowl and shorebirds from wildlife refuges in Alabama, Georgia, and Florida during 2006 to 2008. Swab samples were inoculated into embryonated eggs followed by hemagglutination (HA) test to determine the presence of hemagglutinating agents. Antigen capture-ELISA (AC-ELISA) and real-time reverse transcription-PCR (RRT-PCR) were used to detect AIV from both allantoic fluids (AF) and swab specimens of HA-positive samples. Hemagglutination inhibition test was used to detect Newcastle disease virus, another hemagglutinating virus common in wild birds. The HA-positive AF were sent to the National Veterinary Services Laboratory for subtyping of the isolates. Out of 825 samples tested, 19 AIV and 3 avian paramyxovirus subtypes were identified by the National Veterinary Services Laboratory. Without egg passage, AC-ELISA did not detect virus, whereas matrix gene of 13 AIV were detected using RRT-PCR. When testing was done on AF, 14 were positive for influenza A by AC-ELISA and 20 by RRT-PCR. Antigen capture-ELISA did not detect influenza A when the HA titer was lower than 125, whereas RRT-PCR detected AIV from AF with HA titer as low as 4. The highest isolation rate was from Florida, where out of 109 samples analyzed, 14 AIV were detected by RRT-PCR from AF. Real-time reverse transcription-PCR was more sensitive, specific, and cost-effective than AC-ELISA. However, to avoid false-negative results, testing should be performed on AF and not directly from cloacal swabs. Our procedures to detect AIV directly from cloacal swabs need further optimization for improved sensitivity.

Detection and characterization of avian influenza and other avian paramyxoviruses from wild waterfowl in parts of the southeastern United States.

Cloacal swabs were taken from migratory hunter-killed, nonmigratory, nesting waterfowl and migratory shorebirds from wildlife refuges in Alabama, Georgia, and Florida during 2006 to 2008. Samples were processed in embryonated eggs followed by hemagglutination (HA), Directigen, and real-time reverse transcription-PCR tests. Sequence analysis of the hemagglutinin (H) gene of the H10N7 Alabama isolate revealed that it was closely related (98%) to recent isolates from Delaware and Canada, but only 90% related to an H10N7 isolated 30 yr ago. Four isolates had 94 to 97% similarity to published H1N1 isolates including one from swine. No H5 or H7 isolates were found. One sample was highly pathogenic in embryos, produced a high HA titer, and was positive for both avian influenza (AIV) and Newcastle disease virus or avian paramyxovirus (APMV)-1. In recent (2008) sampling, more (14%) AIV, APMV, or both were isolated than in 2006 to 2007 (1% isolation rate). The higher isolation rate during 2008 may be attributed to optimized sample collection, storage in dry ice, new egg incubator, healthier eggs, time or habitat for isolation, species sampled, migratory status of birds, and more experience with detection procedures. An additional egg passage resulted in increased viral titer; however, no HA-negative samples became HA positive. The chance of transmission of APMV or low-nonpathogenic AIV from wild waterfowl to commercial poultry is possible. However, the chance of transmission of H5 or H7 AIV isolates from waterfowl to commercial farms in Alabama, Georgia, or Florida is unlikely. Therefore, continual testing of these birds is justified to ensure that H5 or H7 AIV are not transmitted to commercial poultry.

Wood duck hatch date: relationship to pairing chronology, plasma luteinizing hormone, and steroid hormones during autumn and winter.

We examined variation in courtship activity and hormone levels of male and female wood ducks (Aix sponsa) in relation to hatch date. Young wood ducks were assigned in groups of 8 (4 males and 4 females) to 4 experimental pens; 2 pens contained early-hatched ducks (3-12 April) and 2 pens contained late-hatched ducks (7-16 June). Courtship behaviors occurred less frequently in October and November than in December and January-February for both early- and late-hatched groups. Early-hatched wood ducks participated in courtship more frequently and formed pair bonds sooner than late-hatched individuals. Testosterone (T) and androstenedione (AD) levels of males did not differ between treatment groups; however, average levels of luteinizing hormone (LH) were greater for early-hatched males. Differences in LH occurred in 2 of 12 samples (6 October and 3 November); otherwise, LH did not vary by treatment. Levels of T and LH in males varied temporally, but there was no significant temporal variation in AE levels. Temporal variation in hormone levels was similar for early- and late-hatched males. Estradiol (E), progesterone (P), and LH levels of females did not differ between treatment groups. There was temporal variation in levels of E and LH, but not of P; this variation was similar for early- and late-hatched females. These data indicate that behavioral differences occurring temporally and between early- and late-hatched wood ducks were not related to corresponding differences in hormone levels.