Skeletal muscle myoblasts are myogenic precursor cells that generate myofibers during muscle development and growth. We recently reported that broiler myoblasts, compared to layer myoblasts, proliferate and differentiate more actively and promptly into myocytes, which corresponds well with the muscle phenotype of broilers. Furthermore, RNA sequencing (RNA-seq) revealed that numerous genes are differentially expressed between layer and broiler myoblasts during myogenic differentiation. Based on the RNA-seq data, we herein report that chicken myoblasts transcribe endogenous retrovirus group K member (ERVK) genes. In total, 16 ERVKs were highly expressed in layer myoblasts and two (termed BrK1 and BrK2) were significantly induced in broiler myoblasts. These transcribed ERVKs had a total of 182 neighboring genes within ±100 kb on the chromosomes, of which 40% were concentrated within ±10 kb of the ERVKs. We further investigated whether the transcription of ERVKs affects the expression of their neighboring genes. BrK1 had two neighboring genes; LOC107052719 was overlapping with BrK1 and downregulated in the broiler myoblasts, and FAM19A2 was upregulated in the broiler myoblasts as well as BrK1. BrK2 had 14 neighboring genes, and only one gene, LOC772243, was differentially expressed between layer and broiler myoblasts. LOC772243 was overlapping with BrK2 and suppressed in the broiler myoblasts. These data indicate that the transcription of ERVKs may impact the expression of their neighboring genes in chicken myoblasts.

We have previously reported a significant association between the single-nucleotide polymorphism (SNP; g.420 C>A) in the cholecystokinin type A receptor gene (CCKAR) and the growth traits of Hinai-dori, a breed of chicken that is indigenous to Japan. Moreover, we have demonstrated that the minor allele of this SNP improved the growth rate in a low-growth line of the Hinai-dori breed. Hence, in the present study, we verified the association between this SNP and the growth traits of the Hinai-jidori chicken: a cross between a Hinai-dori sire and Rhode Island Red dam. In addition, we verified whether the growth rate was improved in Hinai-jidori chickens produced from the parent stocks in which the SNP A/A genotype was fixed by selection (improved Hinai-jidori chickens). The Hinai-jidori female chicks at 4 weeks of age, were subdivided into three genotypic groups (A/A, A/C, and C/C), with 20 chicks in each group, and reared in an open-sided poultry shed until 23 weeks of age. The results showed that the body weight at 23 weeks of age and the average daily gain after 14 weeks of age were significantly higher in group A/A than in group C/C. Subsequently, the improved and the conventional Hinai-jidori chickens were reared until they reached 22 weeks of age to verify the effects on their growth traits. The body weight of the improved Hinai-jidori chickens at 22 weeks was significantly greater than the conventional Hinai-jidori chickens. Moreover, the association between the SNP and body weights of Hinai-jidori chickens at market age (24 weeks) on the production farms showed that the A allele was significantly superior to the C allele. In conclusion, the CCKAR g.420 C>A SNP improves the growth rate of commercial Hinai-jidori chickens and could be a candidate marker for improving the growth performance in selective breeding of Hinai-jidori chickens.

The molecular mechanism underlying photoperiodic response in seasonal breeding animals such as the Japanese quail, red jungle fowl, sheep, mouse, and hamster involves thyroid-stimulating hormone beta subunit (TSHβ) mRNA expression in the pars tuberalis stimulated by the extension in day length. Furthermore, this mechanism is regulated by eyes absent 3 (Eya3) in mammals. Even in birds, the expression of both TSHβ and EYA3 is induced in the pars tuberalis by the extension in day length; however, the relationship between the two genes is unknown. To clarify the function of EYA3 in quail photoperiodism, in the present study, we performed mRNA structure analysis of the Japanese quail EYA3 mRNA using reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot analysis. The results revealed that there are four types of splice variants within regions of exons 7, 8, and 9 of quail EYA3 mRNA. Among the four splice variants of quail EYA3, the splice variant containing exon 7 was expressed in the pars tuberalis on the first long day, when quails were transferred from the short-day condition to the long-day condition. The results indicate that EYA3 splice variant containing exon 7 is involved in the photoperiodic response of the pars tuberalis in the Japanese quail.

We evaluated the effects of supplementing an emulsifier blend (sodium stearoyl-2-lactylate and 1, 3-diacylglycerol) in diets with different energy content (normal and 100 kcal/kg reduced) on the growth performance, meat quality, apparent total tract digestibility (ATTD), and blood lipid profile of broiler chickens. Male broiler chickens (n = 1024), with an initial body weight (BW) of 43.60±0.2 g, were used in a 35-day trial. Broiler chickens of similar body weight were randomly allocated to one of four treatment groups in a 2 × 2 factorial arrangement with two levels of dietary energy content and with or without emulsifier blend. Broiler chickens fed on emulsifier blend supplemented diet had a higher body weight gain (BWG) during d 7–21, d 21–35, and overall period (P<0.05), higher BW during overall period (P<0.05), and lower feed conversion ratio (FCR) during d 7–21, d 21–35, and overall period (P<0.05) compared with broilers fed on diets without emulsifier supplementation. Broiler chickens fed on the diet with low energy content had a lower BWG during d 1–7, d 21–35, and overall period (P<0.05), lower BW during overall period, and higher FCR during d 1–7, d 21–35, and overall period (P<0.05). The ATTD of energy tended to decrease in response to low-energy content diet (P<0.10). Drip loss at 7 d post slaughter tended to decrease in response to dietary emulsifier blend supplementation (P<0.10). However, no interactive effects of dietary energy content and emulsifier blend supplementation (P>0.10) were observed on the growth performance, ATTD, blood lipid profiles, meat quality and relative organ weight. In conclusion, dietary emulsifier blend supplementation could improve growth performance, while low dietary energy content would decrease growth performance and ATTD of energy.

Because of the presence of sperm storage tubules (SSTs) in the utero-vaginal junction (UVJ) in the oviduct, once ejaculated sperm enter the female reproductive tract, they can survive for a prolonged period in domestic birds; however, the specific mechanisms involved in sperm maintenance within the SST remain to be elucidated. In this study, we showed that transferrin (TF) and albumin (ALB) are expressed in SSTs. When UVJ extracts were subjected to size-exclusion column chromatography, we obtained fractions that extend sperm longevity in vitro. LC-MS/MS analysis of the two major proteins in the fractions identified these proteins as TF and ALB. Immunohistochemical analysis using specific antisera against TF and ALB indicated that both proteins were localized not only in the SSTs, but also in the surface epithelium of the UVJ. When the ejaculated sperm were incubated with either purified TF or ALB, sperm viability increased after 24 h. These results indicated that oviductal TF and ALB are involved in the process of sperm storage in SSTs and may open a new approach for technological improvement to prolong sperm longevity in vitro.

We conducted a 28-day trial to evaluate the influence of Bacillus cereus and phytase supplementation on the expression of musculoskeletal strength and intestinal histological features in Japanese quail. Two-hundred day-old quail chicks were divided into four groups with five replicates (n=10): the first group served as a control and was fed only a basal diet (BD); the second group was fed BD + 0.1% B. cereus; the third group was fed BD + 0.01% microbial phytase; and the fourth group was fed BD + 0.01% microbial phytase + 0.1% B. cereus. Compared to the control, individual and combined supplementation of probiotic and phytase increased (P<0.05) the tibial weight, length, outside diameter, and weight of bone ash, but decreased (P<0.05) the weight-to-length index. The water-holding capacity, fiber diameter, fiber cross-sectional area, number of fibers per unit area, fascicle diameter, and fascicle cross-sectional area increased significantly (P<0.05) in birds fed on the combination of phytase and B. cereus. The villus height, width, depth, height-to-crypt depth, and surface area increased significantly (P<0.05) in the B. cereus and phytase groups on an individual basis. The strength of the musculoskeletal system was fully expressed when B. cereus and phytase were given synergistically. However, the histological features of the intestines improved in birds fed on an individual basis.

The goal of this study was to evaluate the influences of mannan oligosaccharides (MOSs) and/or Bifidobacterium on the growth and immunity of pigeons over a 56-day period. One hundred paired adult pigeons were randomly divided into four groups of five paired pigeons. Paired pigeons with two young squabs were housed in a man-made aviary. Parent pigeons in the control group received a basal diet (C), while the other three groups were fed with the basal diet supplemented with 20 g of MOSs/kg of feed (M), 10 g Bifidobacterium (1×1010 CFU/g)/kg of feed (B), or a combination of M and B (MB). We found higher body weights (BW) in pigeons of the B group than in the C, M, and MB groups. None of the treatments exerted significant effects involving spleen and thymus indices, whereas M birds tended to improve the bursa of Fabricius index. Pigeons fed with the M-supplemented diet exhibited improved serum immunoglobulin M (IgM) concentrations compared with those fed with C and the B- and MB-supplemented diets. In addition, M treatment increased immunoglobulin A (IgA) levels compared with MB treatment. MB treatment improved serum immunoglobulin G (IgG) concentrations compared to that by the C treatment. The concentration of secretory immunoglobulin A (sIgA) was significantly reduced in the duodenum and increased in the ileum in pigeons fed with the MB-supplemented diet. This study indicated that dietary supplementation with Bifidobacterium increased the growth performance. Dietary supplementation with MOSs or in combination with Bifidobacterium was able to improve immune function in pigeons but exerted no apparent effect on weight gain. Accordingly, in terms of economic benefits, the findings suggested that supplementation with Bifidobacterium alone may improve production performance, and that supplementation with MOSs alone may improve immune function in pigeons.

Antibiotic growth promoters (AGPs) have been used for many years as supplements in various livestock diets, including those for poultry. However, the use of AGPs in feed was also associated with an increasing number of antibiotic-resistant bacteria in livestock. In this study, the in vitro antibacterial efficacies of eight commercially available non-AGPs suitable for use in poultry were investigated. Assessments included a combination of antibacterial activity assays and estimations of the minimal inhibitory and bactericidal concentrations along with scanning electron microscopy analysis. The results showed that the probiotic, CloStat® exerted a bacteriostatic effect against all tested bacteria, namely Salmonella Typhimurium, Escherichia coli, Staphylococcus aureus, and Clostridium perfringens, whereas Gallipro Tect® and Bacillus Blend® demonstrated bacteriostatic activity towards most of the pathogens tested. Other commercial non-AGPs, Sangrovit®, Fysal®, and Mix oil blend® showed a stronger or equal antibacterial activity compared to the positive control (AGP Maxus® G100) againsts all bacteria tested, except C. perfringens. Nor-Spice AB® and Varium™ did not show any significant effect against the tested bacteria. Several of the tested AGP substitutes exhibited good antibacterial efficiency against pathogenic bacteria and thus may be good candidates for second-stage in vivo investigations into reducing pathogen colonization in broilers.

Glucose is a major circulating carbohydrate in birds and its level in the blood is often used as a biometric indicator in clinical diagnosis and various studies. Notably, hypoglycemia is often associated with Spiking Mortality Syndrome in broilers; therefore, blood glucose levels need to be correctly evaluated in clinical diagnosis. In the present study, we investigated the effect of different blood treatment methods after blood collection on chicken blood glucose measurements. The blood glucose level of plasma separated from blood cell components immediately after blood collection was used as a reference and compared with glucose levels in serum and stored plasma. The mean glucose level in plasma separated from blood cell components immediately after blood collection was 236.1±15.9 mg/dL and remained stable for at least one week in refrigerated storage (between 2°C and 5°C). However, glucose levels decreased slowly in plasma unseparated from blood cell components in storage with ice water. Mean glucose level in serum separated from blood cell components 1 h after blood collection was 206.4±9.2 mg/dL and fell to 108.3±30.0 mg/dL after 24 h. Therefore, the chicken blood serum glucose level was significantly lower than the level in plasma immediately after blood collection, regardless of elapsed time after blood collection. For the measurement of glucose in chicken blood, it is necessary to use refrigeration, use plasma from which blood cell components have been removed, and take measurements within at least 30 min.

Autophagy in the skeletal muscle increases under catabolic conditions resulting in muscle atrophy. This study investigated the effect of inhibition of mechanistic target of rapamycin (mTOR) on autophagy in chick skeletal muscle. We examined the effects of Torin1, an mTOR inhibitor, on autophagy. Chick myotubes were incubated with Torin1 (100 nM) for 3 h. It was observed that Torin1 inhibited the phosphorylation of AKT (Ser473), p70 ribosomal S6 kinase 1 (S6K1, Thr389), S6 ribosomal protein (Ser235/236), and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1, Thr37/46), which are used for measurement of mTOR activity. Torin1 significantly (P< 0.01) increased the LC3-II/LC3-I ratio, an index for autophagosome formation, while it did not influence the expression of autophagy-related genes (LC3B, GABARAPL1, and ATG12). In addition, Torin1 increased atrogin-1/MAFbx (a muscle-specific ubiquitin ligase) mRNA expression. Fasting for 24 h inhibited the phosphorylation of AKT (Ser473), S6K1 (Ther389), S6 ribosomal protein (Ser235/236), and 4E-BP1 (Thr37/46) in chick skeletal muscle and significantly (P<0.01) increased the LC3-II/LC3-I ratio. Fasting also increased GABARAPL1 and atrogin-1/MAFbx mRNA expression but not LC3B or ATG12 mRNA expression. These results indicate that mTOR signaling regulates autophagy and the ubiquitin-proteasome proteolytic pathway in chick skeletal muscle.