Corticosterone in feathers of laying hens: an assay validation for evidence-based assessment of animal welfare.

Studies indicate that the evaluation of animal welfare in birds may be carried out with the measurement of the stress-related hormone corticosterone in feathers. However a standardized procedure for corticosterone measurements in feathers is lacking, a validation needs to be carried out for each new species before implementation. The aim of the present study was to establish a valid method to measure corticosterone concentrations in feathers of laying hens in a precise and repeatable manner using an established and commercially available ELISA. Validation was performed with feather pools of tail and interscapular feathers of commercial Lohmann Brown laying hens. Assessment groups, consisting of 5 replicates, were created. All replicates of an assessment group were processed at the same time. Each replicate was run in 4 repetitions by ELISA. Intra-assay and interassay CV was 7.5 and 6.4%, respectively. The serial dilution showed linearity and parallelism. Examining the hormone extraction efficiency by using different methanol volumes resulted in no statistical differences (P > 0.05). Pulverized feathers showed higher corticosterone values than minced feathers (P > 0.05). Differences were shown between 2 feather types (tail vs. interscapular feathers; P < 0.05), as well as between vane and rachis (P < 0.05). Performance of a freeze-thaw cycle led to a decrease of corticosterone concentrations in the samples. A possible effect of UV-A radiation on the stability of corticosterone in the feathers was not found (P > 0.05). With the present study, a valid protocol, feasible for analyzing feather pools of laying hens, was developed. It may provide fundamentals for further investigations on corticosterone in feathers as a noninvasive indicator to evaluate aspects of animal welfare.

Genetic approach to select against embryo mortality caused by Enterococcus faecalis infection in laying hens.

Selection to reduce susceptibility to Enterococcus faecalis infection in laying hens may contribute to the prevention of amyloid arthropathy since it is mainly induced by this bacterium. Therefore, the aim of the present study was to investigate the possibility to select more laying hens that are resistant against E. faecalis infection through the embryo lethality assay (ELA), which is proposed as an alternative model to replace the adult avian challenge assay. Ten-day-old embryos of 500 Lohmann Brown layers were inoculated into the allantoic cavity with an infectious dose of 2.5 colony-forming units (cfu)/mL of the avian E. faecalis strain K923/96 in 3 ELAs. The embryonic mortality rate (EMR) was determined by candling the eggs daily over a period of 4 d. The average EMR estimated during the 3 ELAs was 50% and the highest EMR occurred 3 to 4 d post inoculation. The estimated heritability for embryonic survivability to the infection was h2 = 0.12-0.14, calculated with the logistic and probit link function, respectively, indicating that the selection of more laying hens that are resistant to E. faecalis infection is feasible. A highly negative genetic correlation was estimated between embryonic survivability and laying performance at the peak of lay (rg = -0.22) and at the end of the production (rg = -0.65), as well as with breaking strength (rg = -0.30 to -0.37). A positive correlation was found between egg weight and the embryonic survivability (rg = +0.16), and no genetic correlation was found with body weight and dynamic stiffness. Therefore, although it was demonstrated that it is possible to breed for E. faecalis resistance, special care should be taken to monitor and to take all traits and their genetic correlations into account in order to achieve a balanced genetic progress. Besides, the ELA does not require that the hens are exposed to pathogens thereby saving labor and cost. Therefore, it could be feasible that ELA be implemented in selection programs.

Reducing the interval of a growth QTL on chromosome 4 in laying hens.

In our previous research, we identified a QTL with an interval of 3.4 Mb for growth on chicken chromosome (GGA) 4 in an advanced intercross population of an initial cross between the New Hampshire inbred line (NHI) and the White Leghorn inbred line (WL77). In the current study, an association analysis was performed in a population of purebred white layers (WLA) with White Leghorn origin. Genotypic data of 130 SNPs within the previously identified 3.4-Mb region were obtained using a 60K SNP chip. In total, 24 significant SNPs (LOD ≥ 4.44) on GGA4 were detected for daily weigh gain from 8 to 14 weeks and two SNPs (LOD ≥ 4.80) for body weight at 14 weeks. The QTL interval was reduced by 1.9 Mb to an interval of 1.5 Mb (74.6-76.1 Mb) that harbors 15 genes. Furthermore, to identify additional loci for chicken growth, a genome-wide association study (GWAS) was carried out in a WLA population. The GWAS identified an additional QTL on GGA6 for body weight at six weeks (19.8-21.2 Mb). Our findings showed that by using a WLA population we were able to further reduce the QTL confidence interval previously detected using a NHI × WL77 advanced intercross population.

Validated quantitative trait loci for eggshell quality in experimental and commercial laying hens.

Compromised eggshell quality causes considerable economic losses for the egg industry. Breeding for improved eggshell quality has been very challenging. Eggshell quality is a trait that would greatly benefit from marker-assisted selection, which would allow the selection of sires for their direct contribution to the trait and would also allow implementation of measurements integrating a number of shell parameters that are difficult to measure. In this study, we selected the most promising autosomal quantitative trait loci (QTL) affecting eggshell quality on chromosomes 2, 3, 6 and 14 from earlier experiments and we extended the F2 population to include 1599 F2 females. The study was repeated on two commercial populations: Lohmann Tierzucht Rhode Island Red line (n = 692 females) and a Hy-Line White Plymouth Rock line (n = 290 progeny tested males). We analyzed the selected autosomal QTL regions on the three populations with SNP markers at 4-13 SNPs/Mb density. QTL for eggshell quality were replicated on all studied regions in the F2 population. New QTL were detected for eggshell color on chromosomes 3 and 6. Marker associations with eggshell quality traits were validated in the tested commercial lines on chromosomes 2, 3 and 6, thus paving the way for marker-assisted selection for improved eggshell quality.

Current approaches to avoid the culling of day-old male chicks in the layer industry, with special reference to spectroscopic methods.

The negative correlation between fattening and laying performance prevents breeding improvement in both laying performance and meat yield. Therefore, specialized chicken lines have been bred in order to achieve either an efficient production of high-quality eggs or high growth rates. As a result, day-old male chicks are culled in the layer hatchery, which poses animal welfare and ethical problems. Breeding companies, scientific groups, and hatcheries are attempting to resolve this issue, with a common aim to find feasible alternatives for the routine killing of male layer chicks. Some approaches aim to influence the sex ratio, while others target at the economically feasible use of the male layer offspring, such as the fattening of "laying hen brothers" or crossbreedings of layers and broilers to create "dual-purpose chickens." Another approach is the sex determination prior to hatch. One of the prerequisites of in ovo sex determination is a practicable method that can be used in industry. The analysis needs to be rapid, cost-efficient, and highly precise; in addition, negative impacts on hatching rate, animal health, and/or performance parameters should be limited. Furthermore, sex determination should be performed before the sensory nervous system's response of the chick embryo to certain or potentially harmful stimuli is developed, which according to current knowledge is before the d 7 of incubation.

Haematological and febrile response to Escherichia coli lipopolysaccharide in 12-week-old cockerels of genetically diverse layer lines fed diets with increasing L-arginine levels.

Due to its decisive function in the avian metabolic, endocrine and immune system L-arginine (Arg) is dietary indispensable for chickens. In 12-week-old cockerels of two high- and two low-performing purebred layer lines, the effects of increasing dietary Arg on the haematological and febrile response were studied over 48 h after single lipopolysaccharide (LPS) injection. The offered diets contained Arg equivalent to 70%, 100% and 200% of recommended supply. Pathophysiological alterations in weight gain, feed intake, body temperature and differential blood count were examined in comparison with their physiological initial values. Within the first 24 h after LPS injection, cockerels reduced feed intake and lost body weight subsequently. Thereby, low-performing genotypes lost body weight to a lesser extent than high-performing ones. The loss of body weight was further intensified by deficient dietary Arg. Within the following 24 h, cockerels recovered by improving feed intake and weight gain. Furthermore, LPS induced genotype-specific fever response: both brown genotypes showed initial hypothermia followed by longer lasting moderate hyperthermia, whereas the white genotypes exhibited biphasic hyperthermia. Fever response was accompanied by significant changes in differential blood counts. Characterized by lymphopenia and heterophilia, a severe leucopenia was observed from 4 to 8 h after LPS injection and replaced by a marked leucocytosis with longer lasting monocytosis up to 48 h after LPS injection. Under given pathophysiological conditions, deficiently Arg-supplied cockerels showed higher total leucocyte counts than adequately and excessively Arg-supplied cockerels. However, deficient and surplus dietary Arg tended to cause higher ratios between heterophils and lymphocytes. To conclude, present results confirmed that LPS induced numerous immunological changes in 12-week-old cockerels and emphasized that chicken's genotype is a source of variation to be considered for immunological studies. Deficient dietary Arg intensified acute changes in differential blood counts and weight gain during LPS-induced inflammation.

Chicken line-dependent mortality after experimental infection with three type IIxIII recombinant Toxoplasma gondii clones.

Three genetically different clones of Toxoplasma gondii, also different in mouse virulence, were studied by experimental infection in chickens. For the experiments, four chicken lines were used, which differed in phylogenetic origin and performance level: two white egg layer lines, one with high laying performance (WLA), one with low (R11) and two brown layer lines, also displaying high (BLA) and low (L68) egg number. Chickens were intraperitoneally infected with three different T. gondii isolates representing type IIxIII recombinant clones, i.e. showing both, type II- and type III-specific alleles. These clones (K119/2 2C10, B136/1 B6H6, K119/2 A7) had exhibited virulence differences in a mouse model. In chickens, a significantly higher mortality was observed in white layer lines, but not in brown layer lines, suggesting that differences in the phylogenetic background may influence the susceptibility of chickens for toxoplasmosis. In addition, antibody (IgY) levels varied in surviving chickens at 31 days post infection. While low to intermediate antibody levels were observed in white layers, intermediate to high levels were measured in brown layers. Infection with a T. gondii clone showing low chicken virulence resulted in higher antibody levels in all chicken lines compared to infection with T. gondii clones of intermediate or high chicken virulence. This was in agreement with the parasite load as determined by real-time PCR. Overall, results show that progeny resulting from natural sexual recombination of T. gondii clonal lineages, may differ in their virulence for mice and chickens.

Metabolic and clinical response to Escherichia coli lipopolysaccharide in layer pullets of different genetic backgrounds supplied with graded dietary L-arginine.

L-arginine (Arg) is an essential amino acid in birds that plays a decisive role in avian protein synthesis and immune response. Effects of graded dietary Arg supply on metabolic and clinical response to Escherichia coli lipopolysaccharide (LPS) were studied over 48 hours after a single intramuscular LPS injection in 18-week-old genetically diverse purebred pullets. LPS induced a genotype-specific fever response within 4 hours post injectionem. Whereas brown genotypes showed an initial hypothermia followed by longer-lasting moderate hyperthermia, white genotypes exhibited a biphasic hyperthermia without initial hypothermia. Furthermore, within 2 hours after LPS injection, sickness behavior characterized by lethargy, anorexia, intensified respiration, and ruffled feathers appeared, persisted for 3 to 5 hours and recovered 12 hours post injectionem. The varying grades of Arg did not alter the examined traits named above, whereas insufficient Arg reduced body growth and increased relative weights of liver and pancreas significantly. At 48 hours post injectionem, increased relative weights of liver and spleen were also found in LPS treated pullets, whereas LPS decreased those of pancreas, bursa, thymus, and cecal tonsils. Moreover, LPS lowered the sum of plasma amino acids and decreased plasma concentrations of Arg, citrulline, glutamate, methionine, ornithine, phenylalanine, proline, tryptophan, and tyrosine, and increased those of aspartate, glutamine, lysine, 1- and 3-methyl-histidine. Elevating concentrations of dietary Arg led to increasing plasma concentrations of Arg, citrulline, ornithine, and 3-methyl-histidine subsequently. As quantitative expression of LPS-induced anorexia, proteolysis, and the following changes in plasma amino acids, pullets showed a significant decrease of feed and nitrogen intake and catabolic metabolism characterized by negative nitrogen balance and body weight loss in the first 24 hours post injectionem. Pullets recovered from the challenge within the second 24 hours post injectionem and changed to anabolism with re-increased feed and nitrogen intake, positive nitrogen retention, and weight gain. To conclude, present results confirmed that LPS induced numerous metabolic and physiological changes in pullet's genotypes, whereas dietary Arg affected the examined traits only slightly.

Implications of genetic selection on yolk proportion on the dry matter content of eggs in a White Leghorn population.

1. The responses to genetic selection on yolk proportion as a technique for increasing egg dry matter content, an important criterion for the egg-product industry, was investigated in a pedigree flock of White Leghorn hens. 2. Parents were preselected on high and low yolk proportion from a base population. The absolute estimated breeding value for yolk proportion of both groups differed by 3%. The realised selection difference in dry matter content of eggs between groups was more than 1% in the analysed offspring population. 3. Heritability estimates were moderate and dry matter had a lower heritability (h(2) = 0.39) than yolk proportion (h(2) = 0.44). 4. The genetic correlation between yolk proportion and dry matter content was highly positive (rg = 0.91). Genetic correlations with egg weight were negative and would have to be compensated for in a breeding programme (rg = -0.76 with yolk proportion and rg = -0.64 with dry matter content). The genetic correlation between the laying performance and yolk proportion was rg = 0.28 and close to zero (rg = -0.05) for dry matter content. 5. Easy recording and lower undesirable correlations make yolk proportion more suitable for commercial selection compared with egg dry matter content in layer breeding.

Genome-wide association study for egg production and quality in layer chickens.

Discovery of genes with large effects on economically important traits has for many years been of interest to breeders. The development of SNP panels which cover the whole genome with high density and, more importantly, that can be genotyped on large numbers of individuals at relatively low cost, has opened new opportunities for genome-wide association studies (GWAS). The objective of this study was to find genomic regions associated with egg production and quality traits in layers using analysis methods developed for the purpose of whole genome prediction. Genotypes on over 4500 birds and phenotypes on over 13,000 hens from eight generations of a brown egg layer line were used. Birds were genotyped with a custom 42K Illumina SNP chip. Recorded traits included two egg production and 11 egg quality traits (puncture score, albumen height, yolk weight and shell colour) at early and late stages of production, as well as body weight and age at first egg. Egg weight was previously analysed by Wolc et al. (2012). The Bayesian whole genome prediction model--BayesB (Meuwissen et al. 2001) was used to locate 1 Mb regions that were most strongly associated with each trait. The posterior probability of a 1 Mb window contributing to genetic variation was used as the criterion for suggesting the presence of a quantitative trait locus (QTL) in that window. Depending upon the trait, from 1 to 7 significant (posterior probability >0.9) 1 Mb regions were found. The largest QTL, a region explaining 32% of genetic variance, was found on chr4 at 78 Mb for body weight but had pleiotropic effects on other traits. For the other traits, the largest effects were much smaller, explaining <7% of genetic variance, with regions on chromosomes 2, 12 and 17 explaining above 5% of genetic variance for albumen height, shell colour and egg production, respectively. In total, 45 of 1043 1 Mb windows were estimated to have a non-zero effect with posterior probability > 0.9 for one or more traits.

Higher precision level at individual laying performance tests in noncage housing systems.

With the Weihenstephan funnel nest box, 12 laying hen flocks were tested for their individual laying performance, egg quality, and nesting behavior in a noncage environment. During the whole observation period of 8 yr, a transponder-based data recording system was continuously improved and resulted in a recording accuracy of 97%. At peak production, heritabilities for the number of eggs laid are in some flocks higher than expected. With improved data accuracy, heritability estimates on individual egg weights are more stable. Heritabilities for nesting behavior traits range between a low to moderate level, providing very useful information for laying hen selection to help improve traits that cannot be recorded in cages. Over the years, the benefits of the Weihenstephan funnel nest box for laying hen breeders have grown. This is due to higher data recording accuracies and extended testing capacities, which result in more reliable genetic parameters.

Enhancing the egg's natural defence against bacterial penetration by increasing cuticle deposition.

The cuticle is a proteinaceous layer covering the avian egg and is believed to form a defence to microorganism ingress. In birds that lay eggs in challenging environments, the cuticle is thicker, suggesting evolutionary pressure; however, in poultry, selection pressure for this trait has been removed because of artificial incubation. This study aimed to quantify cuticle deposition and to estimate its genetic parameters and its role on trans-shell penetration of bacteria. Additionally, cuticle proteins were characterised to establish whether alleles for these genes explained variation in deposition. A novel and reliable quantification was achieved using the difference in reflectance of the egg at 650 nm before and after staining with a specific dye. The heritability of this novel measurement was moderate (0.27), and bacteria penetration was dependent on the natural variation in cuticle deposition. Eggs with the best cuticle were never penetrated by bacteria (P < 0.001). The cuticle proteome consisted of six major proteins. A significant association was found between alleles of one of these protein genes, ovocleidin-116 (MEPE), and cuticle deposition (P = 0.015) and also between alleles of estrogen receptor 1 (ESR1) gene and cuticle deposition (P = 0.008). With the heritability observed, genetic selection should be possible to increase cuticle deposition in commercial poultry, so reducing trans-generational transmission of microorganisms and reversing the lack of selection pressure for this trait during recent domestication.

Analysis of egg production in layer chickens using a random regression model with genomic relationships.

Random regression models allow for analysis of longitudinal data, which together with the use of genomic information are expected to increase accuracy of selection, when compared with analyzing average or total production with pedigree information. The objective of this study was to estimate variance components for egg production over time in a commercial brown egg layer population using genomic relationship information. A random regression reduced animal model with a marker-based relationship matrix was used to estimate genomic breeding values of 3,908 genotyped animals from 6 generations. The first 5 generations were used for training, and predictions were validated in generation 6. Daily egg production up to 46 wk in lay was accumulated into 85,462 biweekly (every 2 wk) records for training, of which 17,570 were recorded on genotyped hens and the remaining on their nongenotyped progeny. The effect of adding additional egg production data of 2,167 nongenotyped sibs of selection candidates [16,037 biweekly (every 2 wk) records] to the training data was also investigated. The model included a 5th order Legendre polynomial nested within hatch-week as fixed effects and random terms for coefficients of quadratic polynomials for genetic and permanent environmental components. Residual variance was assumed heterogeneous among 2-wk periods. Models using pedigree and genomic relationships were compared. Estimates of residual variance were very similar under both models, but the model with genomic relationships resulted in a larger estimate of genetic variance. Heritability estimates increased with age up to mid production and decreased afterward, resulting in an average heritability of 0.20 and 0.33 for pedigree and genomic models. Prediction of total egg number was more accurate with the genomic than with the pedigree-based random regression model (correlation in validation 0.26 vs. 0.16). The genomic model outperformed the pedigree model in most of the 2-wk periods. Thus, results of this study show that random regression reduced animal models can be used in breeding programs using genomic information and can result in substantial improvements in the accuracy of selection for trajectory traits.

Genome-wide association analysis and genetic architecture of egg weight and egg uniformity in layer chickens.

The pioneering work by Professor Soller et al., among others, on the use of genetic markers to analyze quantitative traits has provided opportunities to discover their genetic architecture in livestock by identifying quantitative trait loci (QTL). The recent availability of high-density single nucleotide polymorphism (SNP) panels has advanced such studies by capitalizing on population-wide linkage disequilibrium at positions across the genome. In this study, genomic prediction model Bayes-B was used to identify genomic regions associated with the mean and standard deviation of egg weight at three ages in a commercial brown egg layer line. A total of 24,425 segregating SNPs were evaluated simultaneously using over 2900 genotyped individuals or families. The corresponding phenotypic records were represented as individual measurements or family means from full-sib progeny. A novel approach using the posterior distribution of window variances from the Monte Carlo Markov Chain samples was used to describe genetic architecture and to make statistical inferences about regions with the largest effects. A QTL region on chromosome 4 was found to explain a large proportion of the genetic variance for the mean (30%) and standard deviation (up to 16%) of the weight of eggs laid at specific ages. Additional regions with smaller effects on chromosomes 2, 5, 6, 8, 20, 23, 28 and Z showed suggestive associations with mean egg weight and a region on chromosome 13 with the standard deviation of egg weight at 26-28 weeks of age. The genetic architecture of the analyzed traits was characterized by a limited number of genes or genomic regions with large effects and many regions with small polygenic effects. The region on chromosome 4 can be used to improve both the mean and standard deviation of egg weight by marker-assisted selection.

Genetic variation in eggshell crystal size and orientation is large and these traits are correlated with shell thickness and are associated with eggshell matrix protein markers.

The size and orientation of calcium carbonate crystals influence the structure and strength of the eggshells of chickens. In this study, estimates of heritability were found to be high (0.6) for crystal size and moderate (0.3) for crystal orientation. There was a strong positive correlation (0.65) for crystal size and orientation with the thickness of the shell and, in particular, with the thickness of the mammillary layer. Correlations with shell breaking strength were positive but with a high standard error. This was contrary to expectations, as in man-made materials smaller crystals would be stronger. We believe the results of this study support the hypothesis that the structural organization of shell, and in particular the mammillary layer, is influenced by crystal size and orientation, especially during the initial phase of calcification. Genetic associations for crystal measurements were observed between haplotype blocks or individual markers for a number of eggshell matrix proteins. Ovalbumin and ovotransferrin (LTF) markers for example were associated with crystal size, while ovocleidin-116 and ovocalyxin-32 (RARRES1) markers were associated with crystal orientation. The location of these proteins in the eggshell is consistent with different phases of the shell-formation process. In conclusion, the variability of crystal size, and to a lesser extent orientation, appears to have a large genetic component, and the formation of calcite crystals are intimately related to the ultrastructure of the eggshell. Moreover, this study also provides evidence that proteins in the shell influence the variability of crystal traits and, in turn, the shell's thickness profile. The crystal measurements and/or the associated genetic markers may therefore prove to be useful in selection programs to improve eggshell quality.

Quantitative trait loci affecting eggshell traits in an F(2) population.

Good eggshell quality is important for both table egg quality and chicken reproductive performance. Weak eggshells cause economic losses in all production steps. Poor eggshell quality also poses increased risk for Salmonella infections. Eggshell quality has been a difficult trait to improve by traditional breeding, as it can be measured only for females and it is difficult and expensive to measure. Breeding for improved shell quality may therefore benefit from the use of marker-assisted selection. In an effort to find markers linked to eggshell quality, we have used an F(2) population of 668 females to map quantitative trait loci (QTL) affecting eggshell traits (eggshell deformation, breaking force, weight). By using 160 microsatellite markers on 27 chromosomes, we found 11 genome-wide and 15 suggestive QTL for shell traits measured at different times during production. Loci affecting the deformation were found on chromosomes 1, 2, 6, 10, 14 and Z. Loci affecting the breaking force were detected on chromosomes 2, 3, 10, 12 and Z. Loci affecting the shell weight were detected on chromosomes 6, 12, 24 and Z. Each QTL explains between 1.5% and 4.6% of the phenotypic variance, adding up to 10-15% of total phenotypic variance explained for the different traits. No epistatic effects were observed between loci affecting eggshell traits. Because the effects for quality are mainly additive, these results provide a basis for further characterization of the loci to identify closely linked markers to be used in marker-assisted selection.

Evaluation of inbreeding in laying hens by applying optimum genetic contribution and gene flow theory.

Due to consistent increases of inbreeding of on average 0.95% per generation in layer populations, selection tools should consider both genetic gain and genetic relationships in the long term. The optimum genetic contribution theory using official estimated breeding values for egg production was applied for 3 different lines of a layer breeding program to find the optimal allocations of hens and sires. Constraints in different scenarios encompassed restrictions related to additive genetic relationships, the increase of inbreeding, the number of selected sires and hens, and the number of selected offspring per mating. All these constraints enabled higher genetic gain up to 10.9% at the same level of additive genetic relationships or in lower relationships at the same gain when compared with conventional selection schemes ignoring relationships. Increases of inbreeding and genetic gain were associated with the number of selected sires. For the lowest level of the allowed average relationship at 10%, the optimal number of sires was 70 and the estimated breeding value for egg production of the selected group was 127.9. At the highest relationship constraint (16%), the optimal number of sires decreased to 15, and the average genetic value increased to 139.7. Contributions from selected sires and hens were used to develop specific mating plans to minimize inbreeding in the following generation by applying a simulated annealing algorithm. The additional reduction of average additive genetic relationships for matings was up to 44.9%. An innovative deterministic approach to estimate kinship coefficients between and within defined selection groups based on gene flow theory was applied to compare increases of inbreeding from random matings with layer populations undergoing selection. Large differences in rates of inbreeding were found, and they underline the necessity to establish selection tools controlling long-term relationships. Furthermore, it was suggested to use optimum genetic contribution theory for conservation schemes or, for example, the experimental line in our study.

Breeding to reduce susceptibility to Escherichia coli in layers.

Colibacillosis is a bacterial disease of great concern in the layer industry causing substantial animal and economic losses worldwide. Breeding for resistance to colibacillosis is an important control strategy that would complement traditional management strategies such as vaccinations and therapeutic treatments. Because antibiotic use in animal production is expected to substantially decrease in the future to meet not only consumer demands but also regulations, it is expected that bacterial diseases such as colibacillosis will become even more important. A challenge test was carried out on 353 pedigreed White Leghorn female chickens of 1 pure line. The birds were inoculated with Escherichia coli at an age of 93 to 95 wk (3 hatches) and symptoms of disease and mortality were constantly monitored for up to 6 d after inoculation. The cumulative mortality was 64%, the majority of which took place in the first 2 d. In the present study, genetic parameters of resistance to colibacillosis were estimated to judge the value of genetic selection. The heritability for survival rate was 0.17+/-0.07. Both for quantitative as well as for qualitative traits of economic interest, there was no remarkable difference between survivors and hens that died. Data showed that it is in principle possible to breed for colibacillosis resistance without penalizing the improvement in other key traits. The B21 marker for MHC in this study was associated with a lower colibacillosis susceptibility. In this study, it was demonstrated that it is possible to breed for Escherichia coli resistance. However, due to the inconveniences of a challenge test, it would be very interesting to make use of MAS.

Variation in neighbouring genes of the dopaminergic and serotonergic systems affects feather pecking behaviour of laying hens.

Feather pecking is a behavioural disorder of laying hens and has serious animal welfare and economic implications. One of the several aetiological hypotheses proposes that the disorder results from redirected exploratory behaviour. Variation in the gene encoding the dopamine D4 receptor (DRD4) has been shown to be associated with exploratory behaviour in several species, including in a passerine bird species. We therefore considered DRD4 as a candidate gene for feather pecking. We have annotated DRD4 in the chicken genome and have re-sequenced it in 140 animals belonging to: experimental layer lines divergently selected for high and low propensity to feather pecking; the unselected founder population; and two commercial lines with low and high propensity to feather pecking. We have identified two sub-haplotypes of DRD4 that are highly significantly associated with feather pecking behaviour in the experimental (P = 7.30 x 10(-7)) as well as in the commercial lines (P = 2.78 x 10(-6)). Linkage disequilibrium (LD) extends into a neighbouring gene encoding deformed epidermal autoregulatory factor 1 (DEAF1). The product of DEAF1 regulates the transcription of the gene encoding the serotonin (5-hydroxytryptamine) 1A receptor. Thus, DEAF1 represents another candidate gene for feather pecking. Re-sequencing of five animals homozygous for the 'low-pecking' sub-haplotype and of six animals homozygous for the 'high-pecking' sub-haplotype delineated an LD block of 14 833 bases spanning the two genes. None of the variants in the LD block is obviously functional. However, the haplotype information will be useful to select against the propensity to feather pecking in chicken and to elucidate the functional implications of the variants.

Polymorphisms in eggshell organic matrix genes are associated with eggshell quality measurements in pedigree Rhode Island Red hens.

Novel and traditional eggshell quality measurements were made from up to 2000 commercial pedigree hens for a candidate gene association analysis with organic eggshell matrix genes: ovocleidin-116, osteopontin (SPP1), ovocalyxin-32 (RARRES1), ovotransferrin (LTF), ovalbumin and ovocalyxin-36, as well as key genes in the maintenance and function of the shell gland [estrogen receptor (ESR1) and carbonic anhydrase II (CAII)]. Associations were found for (i) ovalbumin with breaking strength and shell thickness; (ii) ovocleidin-116 with elastic modulus, shell thickness and egg shape; (iii) RARRES1 with mammillary layer thickness; (iv) ESR1 with dynamic stiffness; (v) SPP1 with fracture toughness and (vi) CAII with egg shape. The marker effects are as large as 17% of trait standard deviations and could be used to improve eggshell quality.