The present review focuses on the differential regulation and differential expression, as well as the possible differential functionality, of avian calbindin-D_28k in the three major calcium-transporting tissues: intestine, eggshell gland (ESG) and kidney. Special emphasis is given to their relationships with fast growth and with the laying of long clutches, i.e., unbroken sequences of eggs, separated by one or more days, as affected by the three major regulators, cholecalciferol (vitamin D₃), gonadal activity, and intra- or extracellular calcium or its fluxes. The accumulated evidence suggests that intestinal calbindin mostly depends on vitamin D₃, ESG calbindin appears to depend on calcium-transport-related factor (s) and to lesser extent also on gonadal activity; whereas renal calbindin partly depends on vitamin D₃ and extracellular calcium.

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Individual differentiated somatic cells and undifferentiated stem cells have common genome, although their functions or morphological characters are very different. These differences are derived from difference of gene expression pattern. DNA methylation is generally key factor of suppression of gene and its level is globally change during mammalian early development. But, in birds, whether genome-wide changes in DNA methylation occur during embryonic development is still unknown. Here, we show that genome-wide DNA methylation to assess occurrence during early chick embryonic development. We found that the methylation status at stage 1 was approximately 57%, after which it gradually decreases, reaching a minimum at stage 10 (33%). After stage 10, DNA methylation gradually increased. These results should contribute to clarify the epigenetic mechanisms in birds.

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The present study was conducted to investigate the effects of varying dietary fat levels (3%-10%) on the digestibilities of crude protein, crude fat (CF), nitrogen-free extract and ash at different sites of fistulized chicken intestines. Chickens were fistulized to either the middle part of the jejunum (MJ), the distal end of the jejunum (DJ), the middle part of the ileum, the distal end of the ileum or the distal end of the rectum. Intestinal digesta were collected from each site of intestine, and contents of crude protein, crude fat, nitrogen-free extract and ash were measured. The true digestibility of crude protein in intestinal digesta at MJ in the 10% CF group was significantly lower than that in other groups. The digestibility of crude fat in intestinal digesta at MJ in the 10% CF group was also significantly lower than that in other groups. The digestibility of nitrogen-free extract at MJ and DJ in the 10% CF group was significantly lower than that in other groups. There were no significant differences in digestibility of ash among groups. These results clearly demonstrate that dietary fat levels influence the digestibility of protein, fat, and carbohydrate at MJ in chicken.

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The present study was conducted to evaluate the effect of dietary carnosine on growth performance, carcass characteristics, meat quality, and oxidative stability of muscle and blood in Arbor Acres broilers. A total of 180 1-d-old male broiler chicks were randomly assigned to 3 groups: group A, control feed; group B, 0.50% carnosine feed during d 1 to d 21; group C, 0.50% carnosine feed during d 22 to d 42. Birds were sacrificed at d 42. Live body weight, muscle color (L*, a*, b*), pH values at 0, 2, and 24h postmortem, meat shear force value (SFV), water loss rate, the amount of thiobarbituric acid reactive substances (TBARS), and total antioxidant capability (TAOC) in the blood and the muscle tissue were measured. Results showed that the growth performance of the chickens was not affected by dietary carnosine. The breast muscle weight and thigh muscle weight of group B and the thigh muscle weight of group C was significantly higher than that of group A (P<0.05). The SFV of the breast and the thigh muscle of group C and the SFV of the breast muscle of group B was significantly lower than that of group A (P<0.05). Neither the muscle color nor the pH values at 0, 2, and 24h postmortem were affected. The dietary carnosine decreased the TBARS values and increased the TAOC both in the blood and muscle significantly (P<0.01). These results suggest that dietary carnosine improves chicken meat quantity and quality.

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The study was carried out to investigate the effects of date waste meal (DWM) inclusion in pullet's diet at 0, 4, 8, 12, 16 and 20% and its effect on subsequence performance during laying period. Two hundred and sixteen (16 weeks old) Lohmann Brown pullets were randomly assigned to six experimental groups (3 replicates of 12 pullets/replicate). Pullets were fed a diet containing 16% protein then switched to a 19% protein laying diet until 40 weeks of age.
Results showed that pullets given 20% DWM had earlier sexual maturity by 8 days (P≤0.05) and higher survival rate (95.6%) than the control group. Egg weight was similar among all experimental groups from 20 to 40wks of age. However, higher egg production and better feed conversion ratio were obtained (P≤0.05) at 4% DWM inclusion. Some nutrients digestibility coefficients were significantly improved by feeding 20% DWM. Also, E. coli and fungi counts were reduced (P≤0.05) by about 30.2% and 31.1% as hens received 16 or 20% DWM, respectively as compared to the control group. Total plasma lipids, fresh and stored yolk total lipid concentrations were decreased (P≤0.05) with increasing dietary DWM in diets. A significant increase in egg shell thickness (P≤0.05), yolk index and yolk color was noted as dietary DWM increased to 20%. Haugh units, ovary and oviduct weights percent were not adversely affected by DWM inclusion levels.
In conclusion, DWM could be considered as alternative ingredient for pullet and layer feed. However, more research is needed to determine the optimal DWM inclusion rate.

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Laying performance of Single Comb White Leghorn (SCWL) chicks fed diets containing blood meal supplemented with isoleucine from hatch to ten weeks of age (WOA) was evaluated. In 4 replicates, 480 SCWL chicks were fed corn-based diets whose protein sources were 100% soybean meal (control), 100% blood meal (BM), 50% soybean meal+50% blood meal (SMBM), and 50% alfalfa meal+50% blood meal (AMBM). These diets contained 2,900kcal metabolizable energy (ME) per kg and 20% crude protein (CP). Blood meal comprised 16.8, 11.0 and 14.3% of the dietary composition in BM, SMBM, and AMBM diets, respectively. At 11-14, 15-18 and 19-46WOA birds in all treatment groups were fed corn-soy based diets containing 3,000, 3,080 and 2,900kcal ME/kg and 17.5, 16.5, and 17% CP, respectively. The birds were provided a 23, 8, and 16 hr light regimen at 0-10, 11-14, and 15-46WOA, respectively. Feed and water were provided at free choice and mortality was recorded as it occurred. At first egg, birds were observed for age, body weight and egg weight. Thereafter hen-day egg production, egg weight, egg mass, egg grade, internal egg quality and egg shell thickness were measured over five 28-day laying periods. Birds fed BM diets had 6.08, 5.24 and 2.63% higher (P<0.05) hen-day egg production than those fed the control, AMBM, and SMBM diets, respectively. Mean egg mass of BM fed birds was 5.65, 5.49 and 2.34% higher than that of the control, AMBM, and SMBM birds, respectively. Therefore, feeding diets containing blood meal and supplemented with isoleucine to SCWL chicks at 0-10WOA significantly improved their subsequent egg production performance, but depressed their internal egg quality and egg shell thickness.

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A 4×3 factorial experiment with four dietary fat levels and three protein levels was conducted to determine the influence of dietary fat on performance, egg composition, egg solids, and egg quality of Hy-line W-36 hens at different protein levels. This experiment lasted 16 weeks. Hy-line W-36 hens (n=1440) in phase 1 (21 weeks of age) were randomly divided into 12 treatments (8 replicates of 15 birds per treatment). There was no significant interaction on all parameters between protein and dietary fat. Protein had a significant effect on egg weight, egg mass, feed conversion, albumen weight, percent whole egg solids, and yolk color. Dietary fat had a linear effect on feed intake. As dietary fat increased from 0 to 3.35%, feed conversion linearly improved. A further increase of dietary fat from 3.35 to 5.04% had no additional effect on feed conversion. There was no significant difference in egg specific gravity among hens fed diets supplemented with 0%, 1.67% and 3.35% fat levels. Increasing dietary fat from 3.35 to 5.04% significantly decreased egg specific gravity. As protein increased, albumen weight significantly increased and percent whole egg solids significantly decreased. Increasing dietary fat significantly increased percent yolk, yolk/albumen ratio and percent whole egg solids, but decreased percent albumen. Based on feed conversion and egg specific gravity, the addition of 3.35% fat might be sufficient for optimal performance and egg quality of Hy-line W-36 hens during phase 1.

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We examined the effect of orally administration of taurine on myofibrillar proteolysis in food-deprived chicks. Chicks were food-deprived of 24 hours (h) and then chicks were orally administered 57, 133, or 225mg taurine/100g body weight. Two hours after the administration, the animals were killed and gastrocnemius muscle was taken. Myofibrillar proteolysis was determined based on plasma N^τ-methylhistidine concentrations. Expression of proteolytic-related genes in the muscle was determined by real-time PCR analysis. The administration of taurine decreased myofibrillar proteolysis, as well as the levels of mRNA expression of m-calpain, the C1 and C2 proteasomal subunits, and caspase-3, whereas had no effect on the levels of mRNA expression of ubiquitin, atrogin-1/MAFbx and cathepsin B. These results indicate that oral administration of taurine suppresses myofibrillar proteolysis and the expression of proteolytic-related genes in skeletal muscle of chicks.

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With a view to investigate whether in-ovo cadmium (Cd) exposure can attribute toxic effects in developing avian embryo, the fertile eggs of Japanese quail were injected with Cd and the mortality during incubation and the body weights of day-old hatchlings were measured. The Cd toxicity in the embryo were also assessed by evaluating some well-known oxidative stress markers such as metallothionein (MT) mRNA expression, catalase activity and malondialdehyde (MDA) production in day-10 male and female embryos exposed to Cd with or without ascorbic acid. Results showed that as minimum as 1μg/egg of in-ovo Cd administration increased the embryonic death and decreased the hatchling body weights. A slightly higher mortality rate was found in the 1μg Cd/egg received male embryos than in the female embryos. Compare to the control, higher MT mRNA expression and MDA generation were observed in 1μg Cd/egg received live embryos irrespective of sex. Co-exposure of 50μg ascorbic acid with 1μg Cd/egg inhibited the increse in MDA production in embryos of both sexes but the augment of MT mRNA expression was supressed only in female embryos. These results suggested that in-ovo Cd pollution in avian species might cause the embryonic toxicity leading to death, and the sensitivity of developing quail embryo to antioxidant protection against Cd toxicity is not exactly similar in both sexes of embryos.

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Cellular glutathione peroxidase has important antioxidant and detoxification functions by protecting cells from oxidative damage induced by reactive oxygen species. To investigate the expression of cellular glutathione peroxidase mRNA during embryogenesis in Japanese quail, livers were collected at day 6, 9, 12, 15 and 17 of incubation for measurement of mRNA levels by RT-PCR. The result showed that cellular glutathione peroxidase mRNA expresses in all stages during embryonic development. To observe the effect of selenium in maternal diet on the hatchability and cellular glutathione peroxidase mRNA expression in the liver of hatchlings, mother quail were fed regular diet (0.24ppm selenium) with 0, 0.25, 0.5 or 1ppm selenium. After 2 weeks of feeding, fertilized eggs were collected and incubated. Livers of the mothers and the hatchlings were collected for measurement of cellular glutathione peroxidase mRNA expression. The results showed that the hatchability was high in eggs from quail fed 0.25ppm selenium, but it was decreased in eggs from quail fed 0.5 or 1ppm selenium. Cellular glutathione peroxidase mRNA levels of female quail fed 0.25, 0.5 or 1ppm selenium were higher than those fed regular diet. Cellular glutathione peroxidase mRNA levels of the hatchlings originating from the quail fed 0.25ppm selenium were higher than those of the hatchlings from the quail fed regular diet. These results indicated that although selenium is important for embryonic development through antioxidant defense system by cellular glutathione peroxidase, high level of selenium in the maternal diet has an adverse effect on embryonic development.

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Hinai-dori is a breed of chicken native to Akita Prefecture. The crossbred (Hinai-dori sire×Rhode Island Red) have been commercialized as Hinai-jidori and popular in Japan. Because the taste of male meat is not suitable for the indigenous dish, Kiritanpo stew, Hinai-jidori male has not been used commercially. This paper analyses the effects of caponization at 8 weeks on the meat quality of Hinai-jidori chicken slaughtered at 26 weeks. The thigh and breast meat were used for meat quality analyses, i.e., chemical compositions (moisture, crude protein, and ether extract), meat color, fatty acid composition and histological observations. Caponization caused its meat to be more fat and to decrease redness as compared with males. Caponization resulted in change of the fatty acid profile of thigh meat which was similar to the meat from females. Capon meat became more tender as compared with uncaponized bird and similar in tenderness to female. Regarding muscle structure, it was observed that capons had less connective tissue and thin endomysium as compared with males. These data suggest that caponization improves meat quality and can make unused male chicks usable in the production of Hinai-jidori chickens.

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Hinai-jidori is a popular brand chicken in Japan. Most Hinai-jidori chickens commercially available are females; in contrast, male chicks have been unused. To make unused male Hinai-jidori chicks more usable, an experiment was conducted to examine whether caponization influences growth performance and carcass traits in Hinai-jidori chicken. Hinai-jidori chicks were divided into female, capon, and male groups of 15 birds at 4 weeks of age. Male chicks in the capon group were caponized at 8 weeks of age. Five birds from each of the three groups were slaughtered at 22, 26 and 30 weeks of age, respectively. In the live body weight and daily weight gain, there was no significant difference between the capon and male groups from 18 to 26 weeks of age, while the female group showed the least live body weight and daily weight gain among the three groups throughout the experimental period. In the carcass traits, the capon group showed higher proportion of whole leg and total meat than the female group at 26 and 30 week of age. The capon group had heavier abdominal fat weight than the male group at 26 weeks of age. There was no significant difference in the proportion of abdominal fat between the two groups at all weeks of age. Since capons gain body weight equivalent to males until 26 weeks of age and gain abdominal fat comparable to females, caponization is useful to apply Hinai-jidori males to commercial use.

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Genetically modified cells expressing xenogeneic proteins may trigger strong immunological responses when introduced into animals; however, these responses may be prevented by inducing immunological tolerance. In this study, we have introduced allogeneic cells into chicken embryos via embryonic blood vessel microinjection and induced immunological tolerance to them. Chicken DT40 cells stably transfected with the GFP gene (DT40-GFP cells) were produced and used as antigens to induce immunological tolerance by this method at 65-70h of incubation for preimmunization. Enzyme-linked immunosorbent assays and lymphocyte proliferation assays were performed to determine the serum antibody and lymphocyte responses in hatched chickens after reinjection of the cells at 6 weeks post hatching. Our findings indicated that humoral and cellular immunological responses to allogeneic DT40 cells and DT40-GFP cells expressing xenogeneic proteins can be suppressed by preimmunization during embryogenesis; moreover, an effect on lymphocyte proliferation was observed.

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To investigate whether the genetically modified somatic cells can survive and express foreign gene for a long time after being transplanted into avian embryo, chicken embryonic fibroblasts (CEFs) as a cell vehicle for delivering foreign protein were transfected with the green fluorescent protein (GFP) gene, and were introduced into early chicken embryos via blood vessel microinjection at 65-70h of incubation at 38.5°C. The manipulated eggs were continued to incubate at same condition. The chimerisms of the transplanted CEFs were preliminarily observed under fluorescence microscopy at the different stages in the embryos and the hatchlings. Meanwhile, the chimeric positions of the donor CEFs and the expression of GFP were further examined by polymerase chain reaction (PCR) and immunohistochemistry. The results showed that fluorescent-labeled CEFs embed in the different organs of the recipients including brain, heart, liver, intestine, muscle, etc. and can survive in post-hatch chickens for at least 134 days and the GFP gene can be expressed normally. Long-term survival of the donor CEFs in recipient and normal expression of the GFP gene imply that this approach can be explored for continuous production of target protein in the host chicken.

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Oral infection of Salmonella enterica serovar Enteritidis (SE) strains, SE #15, SE #26, and SE #56, isolated from poultry farms was performed with specific pathogen-free (SPF) 1-day-old chicks and 6 weeks-old BALB/c mice. SE #15 showed the strongest lethality to both chicks and mice, and SE #26 or #56 showed no effects on the survival of mice, while SE #26 showed little and SE #56 intermediate lethal effects to chicks. Using reverse transcriptation-PCR (RT-PCR), expression of pathogenic genes in these SE clones was examined; flagella (fliC), invading host cell (invA), Salmonella enterotoxin (stn) and DNA-binding protein from starved cells of Salmonella (sep22) genes were expressed in SE #15 and SE #56, whereas invA was negative in SE #26. However sep22 was more weakly expressed in SE #26 and SE #56 than SE #15. Western blot analysis of total protein extracts of these SE strains revealed that SE #15 expressed highest levels of SEp22, while SE #26 and SE #56, significantly lower levels of this protein than SE#15. This study showed the extents of sep22 expression at mRNA as well as protein level correlated well with pathogenicity of SE clones to chicks and mice, and these results suggest that SEp22 is a new virulence factor in both chick and mouse.

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