Duration: 10/01/2005 to 09/30/2010

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

Statement of Issues and Justification

The emergence of the US poultry industry during the last half of the 20th century depended upon the following variables: animals characterized by high fecundity, increased performance gained by genetic selection, effective disease prevention, availability of cost-effective feedstuffs, as well as technical advances affecting egg incubation, processing plant operation, and cold storage. Likewise, the success of the US poultry industry is affected by consumer behavior, which in turn is affected by factors such as product cost, health awareness, product diversity, and changing preferences. Most recently, the US poultry industry has become part of a global agribusiness. In summary, the vitality of the contemporary US poultry industry is a function of multiple factors, and many of these are not technical in nature.

Nonetheless, applied science has shown remarkable results. Perhaps the best illustration of this assertion is commercial poultry breeding as practiced over the course of decades. The outcome has been enhanced meat production in record time with improved feed efficiency and greater product uniformity. And yet, this success story is not without problems. For example, the genetic gain realized in turkey growth and body conformation inadvertently resulted in birds that do not mate effectively. This problem has been overcome by use of artificial insemination. Reproductive efficiency in meat-type chickens has also declined as advances in growth have been realized. However, in this case, the setback has not been overcome by application of a simple technology. In fact, this possibility seems unlikely as compromised reproductive efficiency in meat-type chickens appears to stem from a combination of factors that affect gamete production as well as behavior.

As implied above, a commercial poultry industry could never have developed apart from species that produce numerous offspring. Consequently, even though the past and present success of the US poultry industry depends upon many variables, reproduction remains foundational. However, as explained above, genetic selection for growth has compromised reproduction.

The proposed project will build upon the success of a previous regional project directed solely at enhancing the reproductive efficiency of turkeys. A change in scope was necessary for several reasons. First, the number of US poultry scientists with expertise in reproduction has declined in the past decade. This is particularly true for those who work with turkeys. Second, the completion of the chicken genome project affords an unprecedented opportunity to understand reproduction at its most fundamental level: the gene. Nonetheless, though dependent upon genes, reproduction is a process. The collective expertise of scientists involved in this project should facilitate understanding reproductive processes, e.g. photoperiodism, gamete production, or sperm cell function, in terms of the genes that underlie these processes. The third reason a change in scope was warranted is because chicken is the primary type of poultry meat consumed within the US.
A multistate effort is warranted for the purpose of synergy. It is noteworthy that not a single person within this group has a perspective or expertise equivalent to that of the group. This is highly significant because the collective expertise includes knowledge of state-of-the-art research methods, the structure and dynamics of both the primary and secondary breeder industries within the US, the value of technology transfer, and the operation and impact of the US Extension Service. Thus, the collective impact will necessarily be greater than that of any individual. It should also be noted that this group is primarily composed of senior scientists who have an established track record of collaboration, which includes publication of peer-reviewed research, grant writing, and development of novel research tools.
In summary, the proposed project affords a powerful assembly of expertise that addresses a fundamental and pressing problem in a major US agribusiness.

Related, Current and Previous Work

As explained above, the current project is a derivative of a previous regional project entitled "Enhancing the Reproductive Efficiency of Turkeys". The S-285 project was successful in promoting sustained multi-state collaboration. The three objectives were:

1. Determine physiological constraints of gamete and embryo viability.

2. Determine the physiological basis of environmental light control of reproduction.

3. Determine molecular, neuroendocrine and endocrine mechanisms controlling gonadal function.

Objective one was addressed by scientists representing 5 states and the USDA/ARS. Significant accomplishments included the development of the sperm mobility assay and technology transfer at the level of turkey breeders and integrators. Additionally, a greater understanding of incubation conditions, embryonic development and mortality improved our understanding of line differences in development as well as critical survival points for turkey embryos. Variables determining turkey fertility were also extensively studied, providing breeders with useful information on both male and female fertility. This objective was also a popular source of conversation with stakeholders who routinely attended annual meetings.

Objective two was addressed by scientists representing 9 states and the USDA ARS. The projects associated with this objective were instrumental in describing the components and regulation of photoperiodic drive (day-length effects) on the activation and senescence of reproduction in commercial turkeys. Additional work was completed that evaluated the effects of various lighting programs on the persistence of lay in commercial turkeys. Further, collaborative research helped to uncover the relationships between light-dark cycles, biological rhythms, and the ovulatory cycle. This objective was also of interest to stakeholders in that scientists worked on seasonal variation in reproductive performance of commercial birds.

Objective three was addressed by scientists representing 5 states and the USDA/ARS. The projects associated with this objective resulted in the development of both fundamental biology and applied methods for controlling nesting and rearing (broodiness) behavior. The investigators in this project were able to characterize major endocrine components that result in reduced egg production, a critical limiting variable in turkey reproductive efficiency. Vasoactive intestional polypeptide, a small peptide hormone, was found to have profound effects on turkey reproduction. Work was completed to develop an understanding of gene expression associated with reproductive status in the brain, pituitary and gonads. Further work on the development of novel technologies to reduce the impact of reduced egg production on commercial flocks was completed. Stakeholder participation in the application of this knowledge was brisk.

In November 2003, a small group of scientists met in Beltsville, Maryland in order to discuss the development of a new regional project entitled "Enhancing the Reproductive Efficiency of Poultry." The majority of these scientists had been participants in S-285 "Enhancing the Reproductive Efficiency of Turkeys". Critical questions included how the strengths of the previous project could be maintained and yet applied broadly to poultry in general. Three new scientists were recruited: Inma Estevez (Maryland), Dan Satterlee (Louisiana), and Jeanna Wilson (Georgia). As a group, these scientists afforded a new dimension that included expertise in broiler breeder management, animal behavior, endocrinology, and extension as well as additional expertise in the area of technology transfer. The following objectives were proposed.

1. Determine physiological constraints of gamete and embryo viability.

2. Determine physiological and behavioral basis for management of reproduction.

3. Determine molecular, neuroendocrine, and endocrine mechanisms controlling gonadal function.

During the summer of 2004 an open invitation was extended to approximately 20 scientists representing Land Grant universities and the USDA ARS. In October 2004, 15 scientists met in a formal meeting held in Vancouver, Washington. This group decided to move forward with the proposal and at the time of writing, primary participants include Vern Christensen (NCSU), Mohamed El Halawani (Minnesota), Inma Estevez (Maryland), David Froman (OSU), Judy Grizzle (Tennessee), Paul Johnston (BYU), John Kirby (Arkansas), Wayne Kuenzel (Arkansas), Jim Millam (UC Davis), John Proudman (USDA-Beltsville), Ramesh Ramachandran (Penn State), Doug Rhoads (Arkansas), Dan Satterlee (LSU), Tom Siopes (NCSU), Jeanna Wilson (UGA). Additional scientists include Janice Bahr (Illinois), Murray Bakst (USDA-Beltsville), and Julie Long (USDA-Beltsville). The majority of participants are accomplished, senior scientists. Collective expertise includes animal breeding, animal behavior, biorhythm analysis, cell culture, cryopreservation, egg incubation, DNA analysis, gamete biology, endocrinology, embryo physiology, extension, immunocytochemistry, microarray technology, microscopy and imaging, molecular biology, neuroanatomy, neurobiology, neuroendocrinology, proteomics, technology transfer (several participants have a US patent), and toxicology.

Objectives

1. Determine physiological constraints of gamete and embryo viability.
   
2. Determine the physiological and behavioral basis for reproductive management.
   
3. Determine molecular, neuroendocrine and endocrine mechanisms controlling gonadal function.
   

Methods

OBJECTIVE 1: Specific Aim 1: Evaluate the relationship between sperm mobility phenotype and body weight in commercial meat-type chickens. Georgia and Oregon - Sperm mobility is a new quantitative trait in poultry (Froman and Feltmann, 1998). Whereas sperm must be motile to be mobile, motile sperm are not necessarily mobile. Sperm mobility is a primary determinant of male fitness (Froman et al., 1999), and sperm mobility phenotype is subject to genetic selection (Froman and Kirby, 2004). Broiler breeders are selected for growth and feed conversion with little regard for reproductive potential. Consequently, broiler breeder fertility is declining. Preliminary work performed at The University of Georgia (Bowling et al., 2003) has confirmed that broiler breeder males differing in sperm mobility phenotype also differ with respect to fertility, shown that sperm mobility phenotype is independent of testis size as measured by ultrasound, and has shown that males characterized by high sperm mobility may weigh less than low sperm mobility counterparts over the course of a production cycle. In summary, the sperm mobility assay, to date, shows the greatest promise in evaluating the reproductive potential of broiler breeder males. However, an inverse relationship may exist between sperm mobility phenotype and body weight in broiler breeders. This relationship will be re-evaluated on a large scale. Repeated measurements of sperm mobility (Froman et al., 1999) and body weight will be made between 25 to 65 weeks of age using replicate flocks of males. These experiments will afford a comprehensive assessment of how either body weight or rate of gain vary with sperm mobility phenotype in commercial meat-type chickens. Specific Aim 2: Evaluate the effect of mycotoxins on male fitness as measured by sperm mobility and fertility. Tennessee and Oregon - Mycotoxins are highly toxic fungal metabolites that contaminate approximately 25% of the world's food. Ingested mycotoxins induce hepatocelluar carcinoma, immune suppression, kidney damage, hormonal disorders, edema, and compromise reproduction (Prelusky et al., 1994; Leeson et al., 1995). The primary mycotoxins of concern within the US are aflatoxin, T-2 toxin, and diacetoxyscirpenol (DAS; Leeson et al., 1995). Aflatoxins decreased testicular weight and plasma testosterone in both chickens (Sharlin et al., 1980) and quail (Doerr and Ottinger, 1980). Likewise, T-2 toxin induced abnormal sperm development in bobwhites (Grizzle et al., 2002). However, the effect of mycotoxins on sperm mobility is unknown. As explained in the Justification, the fertility of male broiler breeders has been declining for years and this phenomenon has been attributed to intense selection for growth. However, wild bobwhite quail populations have been declining for decades as well (Sisson and Stribling, 2001), and reasons for this decline are unknown. As explained above, sperm mobility is a primary determinant of fertility in male birds (Froman et al., 1999; Birkhead et al., 1999; Donoghue et al., 1999; Donoghue et al., 2003). Therefore, experiments will test the effect of mycotoxins on sperm mobility in broiler breeders and bobwhite quail. Specifically, the effect of single mycotoxins (aflatoxin B1, T-2 toxin, and DAS) will be determined with a dose response using males of predetermined sperm mobility phenotype (Froman et al., 1999) housed in the UT biocontainment facility. Both sperm mobility and fertility will be measured while males are treated with mycotoxins as well as after withdrawal. Thus, acute and chronic effects may be measured. Additionally, tests will be made for an additive effect of low levels of mycotoxins in combination. These experiments will determine if dietary mycotoxins constitute an alternative explanation for compromised male fertility. Specific Aim 3: Evaluate the effect of egg weight on poult quality from breeders maintained at low and high altitude. North Carolina and BYU - Eggshell conductance is a variable that determines embryonic and hatchling livability (Christensen and McCorkle, 1982; Christensen 1983; Ar and Rahn, 1978). Eggshell conductance is dynamic. For example, turkey hens at high altitude lay eggs with reduced conductance in order to conserve water (Rahn et al., 1977, Christensen, unpublished data). Reduced eggshell conductance also prolongs the incubation period and thereby affects poult maturity at hatch (Christensen, unpublished data). The conductance constant for any bird egg is the product of eggshell conductance and incubation period divided by egg weight. Prior studies have examined the effects of eggshell conductance and incubation period on the maturity and livability of poults (Christensen, unpublished data). In contrast, the effect of egg weight has not been addressed when eggshell conductance differs. Therefore, the effect of egg weight on poult quality will be tested using hens from common genetic stock maintained at high altitude (3000 m) and low altitude (100m). Experiments will require approximately 800 eggs for the study of hatchability, 100 embryos for organ analysis (Christensen et al., 2002), and 300 poults for growth trials per low and high altitude flock. Collectively, these data will afford an assessment of poult condition before, during and after hatching relative to egg size. Poults will be grown to market age to test for a latent effect. Experimental variables will include feed consumption, weight gain, and mortality. OBJECTIVE 2: Specific Aim 1: Identify and characterize behavioral components underlying reproductive fitness in poultry. Louisiana and Maryland - Lines of Japanese quail selected for contrasting adrenal stress responsiveness, designated low stress (LS) and high stress (HS), have marked differences in male reproductive secondary sex characteristics and sexual behavior (Satterlee et al., 2002; Jones et al., 2002; Marin and Satterlee, 2003). Selection for reduced adrenocortical responsiveness has accelerated onset of puberty and heightened reproductive potential. This discovery may be relevant to commercial poultry because the Japanese quail is a well-accepted pilot model for extrapolation of research results to species such as chickens and turkeys (Mills and Faure, 1992; Aggrey and Cheng, 1994). However, additional studies are warranted before recommendations can be made pertaining to the overall benefit of selection for reduced adrenal stress responsiveness for poultry in general. Fearful birds are believed to be less active and less social birds are believed to be poor candidates for socio-sexual activity. To date, line differences in fearfulness and sociality are not known to be gender specific. Thus, preference studies will be performed to test for differences in sexual partnering and aggressive behavior. If LS males prefer LS females as opposed to HS or random bred females, line differences will be shown to exist for mating behavior in addition to traits relating to egg and semen production. Related experiments will be performed with chickens. Behavioral studies in red jungle fowl (Lill, 1966; Cheng and Burns, 1988; Collias et al., 1994) and domestic fowl (Jones and Mench, 1991) demonstrated that dominant males have higher reproductive success than subordinates. Such success appears to be due to increased sexual activity of socially dominant chickens (Shabalina, 1984). In support, subordinate chickens were reported to have lower mating frequency (Guhl et al., 1945; Guhl and Warren, 1946; Craig and Bhagwat, 1974). Historically, behaviors such as waltzing and wing flapping frequency as well as physical traits such as comb and wattle size have been associated with dominance status. Recent evidence (McGary et al., 2002, 2003) suggests that certain morphological traits may indeed reflect reproductive success in certain broiler breeder strains. Thus, experiments will be performed to identify fundamental behavioral and morphological traits associated with reduced fertility in yield-type broiler breeders, determine the relationship between these traits and male social status, and evaluate the potential of these traits to be used as markers in genetic selection programs. Likewise, experimentation will address the effect of male location within floor pens on male mating success. Preliminary work (Estevez, unpublished data) demonstrated that highly successful males tend to be distributed toward the center of floor pens. Additional experimentation is warranted using commercial conditions in order to predict how the distribution of socially dominant males affects broiler breeder fertility. In summary, overall fertility of commercial meat-type chickens may be shown to be determined by secondary sexual characteristics and behavior in addition to variables such as body weight or semen quality. Specific Aim 2: Identify and characterize endocrine and physiological components of reproductive fitness in poultry. Louisiana and Arkansas - The hypothalamic-pituitary-testicular (HPT) axis is critical to reproduction. Hypothalamic gonadotropin releasing hormone (GnRH) stimulates secretion of pituitary LH and FSH which, in turn control testis function. LH and FSH bind to receptors on Leydig and Sertoli cells, respectively. LH induces testosterone secretion whereas FSH affects testis development and spermatogenesis (Kirby et al., 1996; Vizcarra et al., 2000). Indeed, a threshold FSH concentration and testis weight have been established at which daily sperm production, i.e. sperm produced per gram testis (DSP), is sufficient for normal sperm production and fertility in broiler breeders (Vizcarra et al., 2000). Male reproduction is compromised when stressors depress the HPT axis. LS and HS lines of quail afford a superb model for studying the means by which adrenal hormones affect the HPT axis. Therefore, line differences will be characterized for circulating levels of steroid and gonadotropin hormones, hypophyseal steady state mRNA FSHb and LHb content, testes weight and morphology, as well as cloacal gland development and function using photostimulated prepubescent, pubertal, young adult, and aged males. Additional variables will include cloacal gland size, testis weight, DSP, and sperm mobility. This study will provide a comprehensive assessment of the relationship between adrenal hormones and the HPT. North Carolina, Minnesota, and USDA Beltsville - This collaboration will attempt to elucidate a precise understanding of how thyroid hormones affect ovarian development and photorefractoriness in turkeys. Whereas photorefractoriness has a profound effect upon fitness in turkey hens, the link between thyroid activity, environmental light, and female reproduction is only beginning to be understood (Siopes, 1997; 2002),. Therefore, the role of thyroid hormones will be determined for the following phenomena in turkey hens: photoinduced ovarian development, the expression of photorefractoriness, and the termination of photorefractoriness. Additionally, phase relationships among thyroid hormones, prolactin, and corticosterone will be examined for the expression and termination of photorefractoriness. Experiments will utilize hormone supplementation in conjunction with light management. In summary, a precise understanding of how thyroid hormones modulate reproduction in the turkey hen may enable practical methods for enhancing reproductive efficiency. Specific Aim 3: Characterization of the role of light on turkey hen reproductive fitness. North Carolina, Minnesota, and UC Davis - As stated previously, photorefractoriness limits fitness in turkey hens. Two general experimental approaches will be used to address this problem. The first involves lighting management. This approach will be used to ask two critical questions. First, when does light initiate or program neural circuits that culminate in prolactin release? Second, can the intensity, wavelength or timing of light exposure be used to attenuate prolactin release thereby keeping hens in lay? In either case, the central variable to be measured is prolactin expression in juvenile females, pubertal females, and adult turkey hens. Pertinent references to this experimental approach include Nichols et al. (1988), Dawson (1991), Wilson and Reinert (1995), Sharp (1996), and Proudman and Siopes (2002). The second experimental approach will address the neuroanatomical and neurochemical basis for photorefractoriness directly. In this case, brain tissue from and female turkeys in various states of photoresponsiveness will be used to identify CNS nuclei, i.e. clusters of related neurons within the central nervous system, gene expression, and neurotransmitter levels that change in response to or following photostimulation. We have recently shown that enkephalinergic axons are apposed with cGnRH-I containing perikarya in the turkey hen's brain (Millam et al., 2002a). Likewise, photoperiod-driven changes in fos expression have been found within the basal tuberal hypothalamus and median eminence of quail Meddle and Follett (1997). In summary, while it understood how prolactin secretion is induced and how turkey hens behave in response to this hormone, the entire neuron network that induces prolactin secretion and how this network is affected by environmental light remains a mystery. The elucidation of the entire informational pathway coupled with a refined knowledge of how light management affects hens will provide a complete model for the biological basis underlying photoperiodic control of egg laying in turkey hens. OBJECTIVE 3: Specific Aim 1: Identify and characterize neuronal circuits controlling reproduction in poultry. Arkansas, USDA Beltsville, and Pennsylvania - Long-day photostimulation that activates the hypothalamo-pituitary-gonadal (HPG) complex is critical for poultry reproduction. GnRH neurons are a key component of this complex. Such neurons are the primary inducers of LH and FSH secretion from the pituitary. However, it is not clear how long-day information is relayed to neurons within the HPG axis. Three sites have been of interest: the eyes, the pineal gland, and the brain (Benoit, 1964; Binkley, 1988; Kuenzel, 1993; Saldanha et al., 2001). To date, the relative contribution of each site unknown, although the encephalic photoreceptors are deemed critical (McMillan et al., 1975; Silver et al., 1988; Wilson, 1991; Kuenzel and Blähser, 1994; Kuenzel et al., 1997). Electrolytic lesioning techniques, immuno-cytochemistry, in situ hybridization histochemistry and imaging procedures will be used to establish which site/s are critical for gonadal function. A primary technique to aid in our analysis of potential photoreceptors is the use of sulfamethazine, a compound that accelerates testis development (Macko Walsh and Kuenzel, 1997). Minnesota, USDA Beltsville, North Carolina, and California - The effort outlined above using chickens will be complemented by two additional studies using turkeys. Previous research has detailed two critical neuroendocrine systems that help control turkey reproduction: the cGnRH I/LH/FSH system and the VIP/prolactin system (Sharp et al., 1998; El Halawani et al., 2001). Two major conceptual leaps have been made that should help outline key nuclei within the brain. First, prolactin secretion is oppositely controlled by dopamine receptors sub-types. Specifically, activation of receptor type D1 stimulates prolactin secretion whereas D2 receptor activation inhibits secretion (Youngren et al., 1995; Chaiseha et al., 1997; Youngren et al., 1998; Chaiseha et al., 2003; Youngren et al., 2002). The second breakthrough was realizing that circulating prolactin is uncoupled from its hypothalamic releasing factor (VIP) in photorefractory hens, i.e. prolactin levels become low even though hypothalamic VIP content is high (Mauro et al., 1992; Saldanha et al., 1994; Chaiseha et al., 1999; Deviche et al., 2000; Teruyama et al., 2001). These insights will aid experimentation that will identify specific hypothalamic nuclei and neuronal phenotypes that have critical roles at three transition points in the turkey reproductive cycle: from reproductive quiescence to recrudescence (photosexual stimulation); from egg-laying to incubation (broodiness); and from the incubation of eggs (broodiness) to brooding chicks. Experiments will use double-label in situ hybridization (Chaiseha et al., 2003) to visualize the retrograde marker, DiI, and mRNAs of critical neuropeptides and rate-limiting enzymes. These include VIP, cGnRH I, tyrosine hydroxylase, and c-fos, a marker of neuronal activation. Related experiments will examine specific actions of VIP on the cGnRH I/LH/FSH system. These experiments will show how VIP influences the behavior of the GnRH/LH/FSH ensemble at key control points (El Halawani et al., 1990; Youngren et al., 1993; Youngren et al., 1994; El Halawani et al., 1996). These experiments will build upon a 20+year history of technical achievement and innovation, including in vitro perfusion of pituitary explants (Chaiseha et al., 1997). The second turkey study will involve imaging techniques to address the neuronal circuitry underlying photosensitive and photorefractory hens. The recently characterized photostimulated c-fos-response of photosensitive hens will be used as a positive control (Millam et al., 2002b). Double-label immunohistochemical and in situ hybridization techniques (Boswell et al., 1998; Millam et al., 2002a) will be employed to determine whether fos-positive neurons collocalize with selected neuropeptides and neurotransmitters that regulate the HPG axis. Likewise, radiolabelled 2-deoxyglucose will be used as a marker of glucose metabolism to compare metabolic activity in brains from photostimulated and photorefractory turkey hens. This approach led to the identification of fos-positive neurons in the tuberal hypothalamus as a neural locus of photorefractoriness (Millam et al., 2002b). This experimental approach is suitable for identifying brain regions that control of reproduction, specifically, the transduction of long daylength cues into cGnRH I outflow in photosensitive hens and the failure of this transduction in photorefractory hens. Specific Aim 2: Identify and characterize regulatory proteins controlling reproduction in poultry. USDA Beltsville, Minnesota, and Arkansas - Proteomics offers great promise for the identification of proteins that differ between two individuals or populations, and is most powerful when applied to discrete cells or groups of cells. USDA-Beltsville has initiated a proteomics program to study cellular factors regulating egg production in the turkey hen. This technology will be used to identify regulatory peptides involved in the two major causes of reproductive failure in the hen: broodiness (incubation behavior) and photorefractoriness. Average egg production of turkey breeder hens ranges from 55 to 100 eggs before they become unprofitable in about 24 weeks of production. However, some hens within a flock will lay 135 eggs in 24 weeks of production, indicating that existing genetic stocks have the potential for much higher egg production. Research over the past two decades by members of this project has helped to identify points at which cellular regulation of reproduction occurs. Members will collaborate on two projects to study how good and poor layers differ at key points of neuroendocrine regulation. Research at Beltsville has shown that a key event in the onset of incubation behaviour is the conversion of pituitary somatotrophs to lactotrophs for the production of prolactin (Ramachandran et al., 1996). At the end of incubation, the added lactotrophs disappear and somatotrophs re-appear, although different cellular mechanisms are employed (Ramachandran et al. 2001). Neural stimulation by eggs or nest on the brood patch has been shown to initiate and maintain broodiness. Our preliminary studies have indicated that nerves from the brood patch synapse in the paraventricular nucleus (PVN) of the brain (Kuenzel et al., 2001). The hypothesis is that incubation behavior is induced and maintained by neural stimulation of neuropeptide(s) synthesized in the PVN and transported to the pituitary via a neural link to the hypothalamus/median eminence and, subsequently, by the hypophyseal portal system, to induce transdifferentiation of somatotrophs to lactotrophs. The proteomes of transdifferentiating pituitaries collected at the onset of incubation and at the termination of incubation will be compared with those of pituitaries from flock mates that do not show broodiness. Proteomes of neurons involved in regulation of prolactin secretion will also be compared. Once the identity of the factor initiating/maintaining transdifferentiation is known, the neural source of this factor will be identified by immunohistochemistry or neural tract tracing. Pennsylvania and USDA Beltsville - Recycling laying hens beyond 72 weeks has economic benefit by reducing egg production cost. Current methods used to induce molt so that lay can be stopped then reinitiated typically involve feed withdrawal. There are two serious drawbacks to this practice. First, it is problematic with respect to animal welfare. Second, feed withdrawal is a stressor that can lead to intestinal infections and subsequent shedding of pathogenic organisms from recycled hens. Thus, alternative methods are sought. The pivotal role of the hypothalamus in reproduction warrants an investigation of the means by which hypothalamic and pituitary function might be manipulated other than by feed withdrawal. The hypothalamus and pituitary gland will be studied at peak lay (32 weeks), at the end of lay (72 weeks), and in response to conventional as well as alternative methods to induce molting. The latter include the use of prolactin, thyroxine, and thyrotropin-releasing hormone-induced. Dependent variables will include gene expression of hypothalamic neuropeptides (cGnRH I, VIP), hypothalamic steroid receptors (progesterone and estrogen), and cell-specific gene expression in gonadotrophs, lactotrophs, folliculo-stellate cells, and somatotrophs within the pituitary. Egg production and eggshell quality will be ascertained to compare the efficacy of various molting treatments. Ovarian follicular hierarchy will be determined to ascertain the reproductive status of each bird. Experiments techniques have been established previously in collaboration with two other participating scientists (Ramachandran et al., 1996; Ramachandran et al., 1998; Ramachandran et al., 2001).

Measurement of Progress and Results

Outputs

• The most readily measured output of the proposed project is the peer-reviewed journal article. While this means of information dissemination has limitations, the peer-reviewed research article remains a primary means by which the effectiveness of a research scientist is measured. Therefore, peer-reviewed journal articles published by participants, i.e. two or more participants as co-authors as opposed to a participant working alone, will be itemized annually in a project report. However, other forms of information sharing include books and book chapters, university-sponsored seminars, professional meetings and symposia, as well as industry-sponsored meetings. Likewise, grantsmanship and technology transfer may also result from collaboration. Therefore, should the project be approved, a means will be developed to document and quantify the effectiveness of collaboration in the broadest sense possible. Additionally, annual meetings will be planned in advance and structured to facilitate information sharing with stakeholders, e.g. representatives of the US primary breeder industry and representatives from companies that manufacture or market reproductive technology.

Outcomes or Projected Impacts

• Improving the fertility of meat-type chickens by understanding how management affects reproductive potential by affecting the hypothalamic-pituitary-gonad axis, which is the critical internal driver of reproduction (Objective 2, Specific Aim 2); by understanding the relationship between sperm mobility, a heritable and highly predictive variable of male fertility, and body weight, a critical end-point for processors (Objective 1, Specific Aim 1); and by understanding how behavior contributes to breeder performance as opposed to sex ratios per se (Objective 2, Specific Aim 1).
• Ascertaining the extent to which mycotoxins, a frequent contaminant of feed, may affect male fitness (Objective 1, Specific Aim 2).
• Optimizing the hatchability of turkey eggs by fully understanding the interrelationship between eggshell conductance, a critical variable that affects embryo livability and poult quality, egg weight, and duration of incubation (Objective 1, Specific Aim 3).
• Understanding the basis for photorefractoriness in turkey hens (Objective 2, Specific Aims 2 and 3; Objective 3, Specific Aim 2).
• Elucidating the neuronal pathway that begins with photoreception and ends with the hypothalamic-pituitary-gonadal axis (Objective 3, Specific Aims 1 and 2).
• Seeking an alternative means for inducing molt by manipulating the hypothalamic-pituitary-gonadal axis by a means other than the stress induced by feed withdrawal (Objective 3, Specific Aim 2).

Milestones

(0):Reducing a long-term decline in the fertility of meat-type chickens.

(0):Developing an adaptive, comprehensive understanding of turkey egg hatchability that can be applied at the level of commercial hatcheries.

(0): Discovering the basis for photorefractoriness in turkeys.

(0): Explaining CNS integration that drives the hypothalamic-pituitary-gonadal axis.

Projected Participation

View Appendix E: Participation

Outreach Plan

As explained above, annual meetings will be planned in advance and structured to facilitate information sharing with stakeholders, e.g. representatives of the US primary breeder industry and representatives from companies that manufacture or market reproductive technology. It is noteworthy that several participants have FTE in the Extension Service and track records as accomplished faculty in this mission of the Land Grant university. It is also noteworthy that most of the participants have working relationships with key people at businesses such as Animal Reproduction Systems, BUTA, Cobb-Vantress, Hybrid Turkeys, Nicholas Farms, Perdue Farms, etc. Therefore, any emergent outreach plan will be determined and enacted by the group rather than the responsibility of any given member.

Organization/Governance

Chair: David Froman, Oregon State University (David.Froman@oregonstate.edu)

Secretary: Doug Rhoads, University of Arkansas.

Each annual meeting will conclude with a formal business meeting.

Literature Cited

Aggrey, S.E., and K.M. Cheng, 1994. Animal model analysis of genetic (Co) variances for growth traits in Japanese quail. Poultry Sci. 73: 1822-1828.

Ar, A., and H. Rahn, 1978. Interdependence of gas conductance, incubation length and weight of the avian egg. Pages 227-236 In: Respiratory Function in Birds, Adult and Embryonic, J. Piiper, ed., Springer-Verlag, London.

Benoit, J. 1964. The role of the eye and the hypothalamus in photostimulation of gonads in the duck. Ann. N.Y. Acad. Sci. 117:204-217.

Binkley, S. 1988. The pineal. Prentice-Hall, Englewood Cliffs, N.J.

Birkhead, T.R., J.G. Martinez, T. Burke, and D.P. Froman, 1999. Sperm mobility determines the outcome of sperm competition in the domestic fowl. Proc. R. Soc. Lond. B 266:1759-1764.

Bowling, E.R., D.P. Froman, A.J. Davis, and J.L. Wilson, 2003. Attributes of broiler breeder males characterized by low and high sperm mobility. Poultry Sci. 82:1796-1801.

Chaiseha, Y., O.M. Youngren, and M.E. El Halawani, 1997. Dopamine receptors influence vasoactive intestinal peptide release from turkey hypothalamic explants. Neuroendocrinology.65: 423-429.

Chaiseha,Y., O.M. Youngren, and M.E. El Halawani, 1997. Dopamine receptors influence vasoactive intestinal peptide release from turkey hypothalamic explants. Neuroendocrinology 5: 423-429.

Chaiseha, Y. and M.E. El Halawani, 1999. Expression of vasoactive intestinal peptide/peptide histidine isoleucine in several hypothalamic areas during the turkey reproductive cycle in relationship to prolactin secretion. Neuroendocrinology 70:402-412.

Chaiseha, Y., O.M. Youngren, K. Al-Zailaie, and M.E. El Halawani, 2003. Expression of D1 and D2 dopamine receptors in the hypothalamus and pituitary during the turkey reproductive cycle: colocalization with vasoactive intestinal peptide. Neuroendocrinology 77:105-118.

Cheng, K. M., and J. T. Burns. (1988). Dominance relationship and mating behavior of domestic cocks - a model to study mate-guarding and sperm competition in birds. The Condor 90: 697-704.

Christensen , V.L., 1983. Distribution of pores on hatching and nonhatching turkey eggs. Poultry Sci. 62:1312-1316.

Christensen, V.L., and F.M. McCorkle, 1982. Turkey egg weight losses and embryonic mortality during incubation. Poultry Sci. 61:1209-1213.

Christensen, V.L., M.J. Wineland, G.M. Fasenko, and W.E. Donaldson, 2002. Egg storage alters weight of supply and demand organs of broiler chicken embryos. Poultry Sci. 81:1738-1743.

Collias, N., E. Collias, and R.I. Jennrich, 1994. Dominant red jungle fowl (Gallus gallus) hens in an unconfined flock rear the most young over their lifetime. Auk 111: 863-872.

Craig, J.V., and A.L. Bhagwat, 1974. Agonistic and mating behavior of adult chickens modified by social and physical environments. Appl. Anim. Ethology 1(1): 57-65.

Dawson, A., 1991. The induction of photorefractoriness and molt in starlings, Sturnis vulgaris, by continuous or intermittent long days. Physiol. Zool. 64: 1252-1261.

Deviche, P., C.J. Saldanha, and R. Silver, 2000. Changes in brain gonadotropin-releasing hormone- and vasoactive intestinal polypeptide-like immunoreactivity accompanying reestablishment of photosensitivity in male dark-eyed junco (Junco hyemalis). Gen. Comp. Endocrinol.117: 81-97.

Doerr, J.A. and M.A. Ottinger. 1980. Delayed reproductive development resulting from aflatoxicosis in juvenile Japanese quail. Poultry Sci. 59:1995-2001.

Donoghue, A.M., T.S. Sonstegard, L.M. King, E.J. Smith, and D.W. Burt, 1999. Turkey sperm mobility influences paternity in the context of competitive fertilization. Biol. Reprod. 61:422-427.

Donoghue, A.M., J.D. Kirby, D.P. Froman, S.P. Lerner, A.N. Crouch, L.M. King, D.J. Donoghue, and T.S. Sonstegard, 2003. Field testing the influence of sperm competition based on sperm mobility in breeder turkey toms. Brit. Poult. Sci. 44:1-7.

El Halawani, M,E., S.L. Silsby, and L.J. Mauro, 1990. Vasoactive intestinal peptide is a hypothalamic prolactin-releasing neuropeptide in the turkey (Meleagris gallopavo). Gen. Comp. Endocrinol. 78: 66-73.

El Halawani, M.E., O.M. Youngren, and Y. Chaiseha, 2001. Neuroendocrinology of prolactin regulation in the domestic turkey, in Dawson M, Chaturvedi CM (eds): Avian Endocrinology. Narosa Publishing House, New Delhi, India, pp 233-244.

Froman, D.P., and A.J. Feltmann, 1998. Sperm mobility: A quantitative trait of the domestic fowl (Gallus domesticus). Biol. Reprod. 58:379-384.

Froman, D.P., and J.D. Kirby, 2004. Sperm mobility: Phenotype in roosters (Gallus domesticus) determined by mitochondrial function. Biol. Reprod. (in press).

Froman, D.P., A.J. Feltmann, M.L. Rhoads, and J.D. Kirby, 1999. Sperm mobility: A primary determinant of fertility in the domestic fowl (Gallus domesticus). Biol. Reprod. 61:400-405.

Grizzle, J.M., D. McCracken, D. Kersten, C. Hernandez, A. Houson, and A. Saxton, 2002. T-2 mycotoxicosis in bobwhite quail. http://www.agriculture.utk.edu/ansci/pdf/Reports/t2mycotoxicosis.pdf

Guhl, A.M., N.E. Collias, and W.C. Allee, 1945. Mating behavior and the social hierarchy in small flocks of white leghorns. Physiol. Zool. 18: 365390.

Guhl, A.M., and D.C. Warren, 1946. Number of offspring sired by cockerels related to social dominance in chickens. Poultry Sci. 25: 460472.

Jones, M.E.J., and J.A. Mench, 1991. Behavioral correlates of male mating success in a multisire flock as determined by DNA fingerprinting. Poultry Sci. 70: 1493-1498.

Jones, R.B., R.H. Marin, D.G. Satterlee, and G.G. Cadd, 2002. Sociality in Japanese quail (Coturnix japonica) genetically selected for contrasting adrenocortical responsiveness. Appl. Anim. Behav. Sci. 75: 337-346.

Kirby, J.D., M.V. Mankar, D. Hardesty, and D.L. Kreider, 1996. Effects of transient prepubertal 6-N-propyl-2-thiouracil treatment on testis development and function in the domestic fowl. Biol. Reprod. 55: 910-916.

Kuenzel, W.J., 1993. The search for deep encephalic photoreceptors within the avian brain, using gonadal development as a primary indicator. Poultry Sci. 72:959-967.

Kuenzel, W.J. and S. Blähser, 1994. Vasoactive intestinal polypeptide (VIP)-containing neurons: distribution throughout the brain of the chick (Gallus domesticus), with focus upon the lateral septal organ. Cell Tissue Res. 275:91-107.

Kuenzel, W.J., S.K. McCune, R.T. Talbot, P.J. Sharp and J.M. Hill, 1997. Sites of gene expression for vasoactive intestinal polypeptide throughout the brain of the chick (Gallus domesticus). J. Comp. Neur. 381:101-118.

Kuenzel, W.J., R. Ramesh, J.A. Proudman, and R.R. Miselis, 2001. A potential neural tract-tracing method for use in avian species. Poultry Sci. 80(Suppl. 1):176.

Leeson, S., G.J. Diaz, and J. D. Summers, J.D., 1995. Trichothecenes, In: Poultry metabolic disorders and mycotoxins. University Books, Guelph, Ontario, pp. 190-226.

Lill, A., 1966. Some observations on social organization and non-random mating in captive Burmese red jungle fowl (Gallus gallus spadiceus). Behaviour 26: 228-241.

Macko Walsh, K. and W.J. Kuenzel, 1997. Effect of sulfamethazine on sexual precocity and neuropeptide Y neurons within the tuberoinfundibular region of the chick brain. Brain Res. Bull. 44:707-713.

Marin, R.H. and D.G. Satterlee, 2003. Selection for contrasting adrenocortical responsiveness in Japanese quail (Coturnix japonica) influences sexual behavior in males. Appl. Anim. Behav. Sci. 83: 187-199.

Mauro, L.J., Youngren, O.M., Proudman, J.A., Phillips, R.E. and El Halawani, M.E., 1992. Effects of reproductive status, ovariectomy, and photoperiod on vasoactive intestinal peptide in the female turkey hypothalamus. Gen. Comp. Endocrinol. 87:481-493.

McGary, S., I. Estévez, M. Bakst, and D. Pollock, 2002. Phenotypic traits as reliable indicators of fertility in male broiler breeders. Poultry Sci. 81:102-111.

McGary, S., I. Estévez and M. Bakst, 2003. Potential relationships between physical traits and male broiler breeder fertility. Poultry Sci. 82:328-337.

McMillan, J.P., H.A. Underwood, J.A. Elliott, M.H. Stetson and M. Menaker, 1975. Extraretinal light perception in the sparrow. IV. Further evidence that the eyes do not participate in photoperiodic photoreception. J. Comp. Physiol. 97:205-213.

Meddle, S.L., and B.K. Follett, 1997. Photoperiodically driven changes in fos expression within the basal tuberal hypothalamus and median eminence of Japanese quail. J. Neuroscience 17: 8909-8918.

Millam, J.R., R. Wang, C.B. Craig-Veit, and T.D. Siopes. 2002a. Apposition of enkephalinergic axons with cGnRH I-containing perikarya in turkey hen brain. Gen. Comp. Endocrinol. 127: 128-135.

Millam, J.R., C.B. Craig-Veit, and T.D. Siopes, 2002b. Photostimulated fos-like immunoreactivity in tuberal hypothalamus of photosensitive vs. photorefractory turkey hens. Poultry Sci. 81(Suppl. 1): 79.

Mills, A.D., and J.M Faure, 1992. The behaviour of domestic quail. In: Nutztierethologie. M. Nichelmann, Ed. Gustav Fisher Verlag, Jena, Germany, pp. 1-16.

Nicholls, T.J., A.R. Goldsmith, and A. Dawson, 1988. Photorefractoriness in birds and in comparison with mammals. Physiol. Rev. 68:133-176.

Prelusky, D.B., B.A. Rotter, and R.G. Rotter, 1994. Toxicology of mycotoxins. In: Miller, J. D. and Trenholm, H. L., eds. Mycotoxins in Grain, compounds other than aflatoxins. Eagen Press, St. Paul, Minnesota, USA, pp. 359-405.

Proudman, J. A., and T. D. Siopes, 2002. Relative and absolute photorefractoriness in turkey hens: Profiles of prolactin, thyroxine, and triiodothyronine early in the reproductive cycle. Poultry Sci. 81: 1218-1223.

Rahn, H., C. Carey, K. Balmas, B. Bhatia, and C.V. Paganelli, 1977. Reduction of pore area of the avian eggshell as an adaptation to altitude. Proc. Nat. Acad. Sci. 74:2095-2098.

Ramachandran, R., J.A. Proudman, and W.J. Kuenzel, 1996. Changes in pituitary somatotroph and lactotroph distribution in laying and incubating turkey hens. Gen. Comp. Endocrinol. 104:67-75.

Ramachandran, R., R. Solow, J.A. Proudman, and W.J. Kuenzel, 1998. Identification of mammosomatotrophs in the turkey hen pituitary: Increased abundance during hyperprolactinemia. Endocrinology 139:781-786.

Ramachandran, R., W.J. Kuenzel, and J.A. Proudman, 2001. Increased proliferative activity and programmed cellular death in the turkey hen pituitary gland following interruption of incubation behavior. Biol. Reprod. 64:611-618.

Saldanha, C.J., P.J. Deviche, and R. Silver, 1994. Increased VIP and decreased GnRH expression in photorefractory dark-eyed Juncos (Junco hyemalis). Gen. Comp. Endocrinol. 93:128-136.

Saldanha, C.J., A.-J. Silverman and R. Silver, 2001. Direct innervation of GnRH neurons by encephalic photoreceptors in birds. J. Biol. Rhythms 16:39-49.

Satterlee, D.G., R.H. Marin, and R.B Jones, 2002. Selection of Japanese quail for reduced adrenocortical responsiveness accelerates puberty in males. Poultry Sci. 81: 10711076.

Shabalina, A.T., 1984. Dominance rank, fear scores and reproduction in cockerels. Brit. Poultry Sci. 25: 297-301.

Sharlin, J.S., B. Howarth, Jr., and R.D. Wyatt. 1980. Effect of dietary aflatoxin on reproductive performance of mature White Leghorn males. Poultry Sci. 59:1311-1315.

Sharp, P. J., 1996. Strategies in avian breeding cycles. Anim. Reprod. Sci. 42: 505-513.

Sharp, P.J., A. Dawson, and R.W. Lea, 1998. Control of luteinizing hormone and prolactin secretion in birds. Comp. Biochem. Physiol. C Pharmacol. Toxicol Endocrinol. 119:275-82.

Siopes, T., 1997. Transient hypothyroidism reinitiates egg laying in turkey breeder hens: termination of photorefractoriness by propylthiouracil. Poultry Sci. 76: 1776-1782.

Siopes, T., 2002. Circulating thyroid hormone levels in recycled turkey breeder hens during a short day prelighting period and renewal of photosensitivity for egg production. Poultry Sci. 81: 1342-1346.

Silver, R., P. Witovsky, P. Horvath, V. Alones, C.J. Barnstable and M.N. Lehman, 1988. Coexpression of opsin-and VIP-like-immunoreactivity in CSF-contacting neurons of the avian brain. Cell Tissue Res. 253:189-198.

Sisson, C., and L. Stribling, 2001. The, rise, fall, and resurrection of the bobwhite quail in the Southeast. Wildlife Trends. 1:5-8.

Teruyama, R., and M.M. Beck, 2001. Double immunocytochemistry of vasoactive intestinal peptide and cGnRH-I in male quail: photoperiodic effects. Cell Tissue Res.303: 403-414.

Vizcarra, J.A., W.L. Bacon, and J.D. Kirby, 2000. Physiological factors affecting the reproductive performance of commercial broiler breeder males. Proc. 49th Ann. Natl. Breeders Roundtable, St. Louis, MO, pp. 18-22.

Wilson, F.E., 1991. Neither retinal nor pineal photoreceptors mediate photoperiodic control of seasonal reproduction in American Tree sparrows (Spizella arborea). J. Exptl Zool. 259:117-127.

Wilson, F.E., and B.D. Reinert, 1995. The photoperiodic control circuit in euthyroid American tree sparrows is already programmed for photorefractoriness by week four under long days. J. Reprod. fertil. 103: 279-284.

Youngren, O.M., M.E. El Halawani, J.L. Silsby, and R.E. Phillips, 1993. Effect of reproductive condition on luteinizing hormone and prolactin release induced by electrical stimulation of the turkey hypothalamus. Gen. Comp. Endocrinol. 89: 220-228.

Youngren, O.M., J.L. Silsby, I. Rozenboim, R.E. Phillips, and M.E. El Halawani, 1994. Active immunization with vasoactive intestinal peptide prevents the secretion of prolactin induced by electrical stimulation of the turkey hypothalamus. Gen. Comp. Endocrinol.95: 330-336.

Youngren, O.M., G.R. Pitts, R.E. Phillips, and M.E. El Halawani, 1995. The stimulatory and inhibitory effects of dopamine on prolactin secretion in the turkey. Gen. Comp. Endocrinol. 98:111-117.

Youngren, O.M., Y. Chaiseha, and M.E. El Halawani, 1998. Regulation of prolactin secretion by dopamine and vasoactive intestinal peptide at the level of the pituitary in the turkey. Neuroendocrinology 68: 319-325.

Youngren, O., Y. Chaiseha, K. Al-Zailaie, S. Whiting, S. Kang, S., and M.E. El Halawani, 2002. Regulation of prolactin secretion by dopamine at the level of the hypothalamus in the turkey. Neuroendocrinology 75: 185-192.

Attachments

Land Grant Participating States/Institutions

AL, AR, CA, GA, IL, LA, MD, MN, NY, OR, PA, TN

Non Land Grant Participating States/Institutions

Spain, USDA-ARS