Research Note: Prospects for early detection of breast muscle myopathies by automated image analysis.

White Striping (WS), Wooden Breast (WB), and Spaghetti Meat (SM) are documented breast muscle myopathies (BMM) affecting broiler chickens' product quality, profitability and welfare. This study evaluated the efficacy of our newly developed deep learning-based automated image analysis tool for early detection of morphometric parameters related to BMM in broiler chickens. Male chicks were utilized, and muscle samples were collected on d 14 of rearing. Histological procedures, including microscopic scoring, blood vessel count, and collagen quantification, were conducted. A previous study demonstrated our automated image analysis as a reliable tool for evaluating myofiber size, conforming with manual histological measurements. A threshold for BMM detection was established by normalizing and consolidating myofiber diameter and area into a unified metric based on automated measurements, also termed as "relative myofiber size value." Results show that severe myopathy broilers consistently exhibited lower relative myofiber size values, effectively detecting myopathy severity. Our study, aimed as proof of concept, underscores the potential of our automated image analysis tool as an early detection method for BMM.

In-ovo feeding with creatine monohydrate: implications for chicken energy reserves and breast muscle development during the pre-post hatching period.

The most dynamic period throughout the lifespan of broiler chickens is the pre-post-hatching period, entailing profound effects on their energy status, survival rate, body weight, and muscle growth. Given the significance of this pivotal period, we evaluated the effect of in-ovo feeding (IOF) with creatine monohydrate on late-term embryos' and hatchlings' energy reserves and post-hatch breast muscle development. The results demonstrate that IOF with creatine elevates the levels of high-energy-value molecules (creatine and glycogen) in the liver, breast muscle and yolk sac tissues 48 h post IOF, on embryonic day 19 (p < 0.03). Despite this evidence, using a novel automated image analysis tool on day 14 post-hatch, we found a significantly higher number of myofibers with lower diameter and area in the IOF creatine group compared to the control and IOF NaCl groups (p < 0.004). Gene expression analysis, at hatch, revealed that IOF creatine group had significantly higher expression levels of myogenin (MYOG) and insulin-like growth factor 1 (IGF1), related to differentiation of myogenic cells (p < 0.01), and lower expression of myogenic differentiation protein 1 (MyoD), related to their proliferation (p < 0.04). These results imply a possible effect of IOF with creatine on breast muscle development through differential expression of genes involved in myogenic proliferation and differentiation. The findings provide valuable insights into the potential of pre-hatch enrichment with creatine in modulating post-hatch muscle growth and development.

Vertical transmission of gut bacteria in commercial chickens is limited.

The existence of vertical transmission in chickens under commercial settings, where chicks are raised separately from adults, is unclear. To answer this question, the fecal microbiota of chicks hatched and grown separately was compared with their mothers' microbiota. Most amplicon sequence variants (ASVs) identified in hens were not detected at all in chicks up to two weeks of age by 16S rDNA sequencing, and those that were detected had a low incidence among the chicks. Nevertheless, a few ASVs that were common with the hens were highly prevalent among the chicks, implying that they were efficiently transmitted to chicks. These ASVs were culturable from the reproductive tract of hens and eggshells. Furthermore, interventions attempting to disrupt transmission resulted in a reduction in the prevalence of specific phylogenetic groups in chicks. To conclude, vertical transmission in commercial poultry grown separately from adults likely exists but is not efficient, possibly resulting in impairment of microbiota function. This implies that artificial exposure to adult bacterial strains might improve microbiota functioning.

A deep learning-based automated image analysis for histological evaluation of broiler pectoral muscle.

Global market demand for chicken breast muscle with high yield and quality, together with the high incidence rate of breast muscle abnormalities in recent years highlights the need for tools that can provide a rapid and precise evaluation of breast muscle development and morphology. In this study, we used a novel deep learning-based automated image analysis workflow combining Fiji (ImageJ) with Cellpose and MorphoLibJ plugins to generate an automated diameter and cross-sectional area quantification for broiler breast muscle. We compared data of myofiber diameter from 14-day-old broiler chicks, generated either by manual analysis or by automated analysis. Comparison between manual and automated analysis methods exhibited a striking accuracy rate of up to 99.91%. Moreover, the automated analysis method was much faster. When the automated analysis method was implemented on 84 breast muscle cross-section images it characterized 59,128 myofibers within 4.2 h, while manual analysis of 27 breast muscle cross-section images enabled analysis of 17,333 myofibers in 54 h. The automated image analysis method was also more productive, producing data sets of both diameter and cross-sectional area at an 80-fold higher rate than the manual analysis (26,279 vs. 321 data sets per hour, respectively). In order to demonstrate the ability of this automated image analysis tool to detect differences in breast muscle histomorphology, we applied it on cross sections from chicks of control and in ovo feeding group, injected with a methionine source [2-hydroxy-4-(methylthio) butanoic calcium salt (HMTBa)], known to effect skeletal muscle histomorphology. Analysis was performed on 19,807 myofibers from the control group and 21,755 myofibers from the HMTBa group and was completed in less than 1 h. The clear advantages of this automated image analysis workflow characterized by high precision, high speed, and high productiveness demonstrate its potential to be implemented as a reproducible and readily adaptable research or diagnostic tool for chicken breast muscle development and morphology.

Profiling intestinal stem and proliferative cells in the small intestine of broiler chickens via in situ hybridization during the peri-hatch period.

Mature small intestines have crypts populated by stem cells which produce replacement cells to maintain the absorptive villus surface area. The embryonic crypt is rudimentary and cells along the villi are capable of proliferation. By 7 d post-hatch the crypts are developed and are the primary sites of proliferation. Research characterizing the proliferative expansion of the small intestine during the peri-hatch period is lacking. The objective of this study was to profile the changes of genes that are markers of stem cells and proliferation: Olfactomedin 4 (Olfm4), Leucine-rich repeat containing G protein-coupled receptor 5 (Lgr5), and marker of proliferation Ki67 from embryonic day 17 to 7 d post-hatch using quantitative PCR and in situ hybridization (ISH). The expression of the stem cell marker genes differed. Olfm4 mRNA increased while Lgr5 mRNA decreased post-hatch. Ki67 mRNA decreased post-hatch in the duodenum and was generally the greatest in the ileum. The ISH was consistent with the quantitative PCR results. Olfm4 mRNA was only seen in the crypts and increased with morphological development of the crypts. In contrast Lgr5 mRNA was expressed in the crypt and the villi in the embryonic periods but became restricted to the intestinal crypt during the post-hatch period. Ki67 mRNA was expressed throughout the intestine pre-hatch, but then expression became restricted to the crypt and the center of the villi. The ontogeny of Olfm4, Lgr5, and Ki67 expressing cells show that proliferation in the peri-hatch intestine changes from along the entire villi to being restricted within the crypts.

Supply and demand of creatine and glycogen in broiler chicken embryos.

Optimal embryonic development and growth of meat-type chickens (broilers) rely on incubation conditions (oxygen, heat, and humidity), on nutrients and on energy resources within the egg. Throughout incubation and according to the embryo's energy balance, the main energy storage molecules (creatine and glycogen) are continuously utilized and synthesized, mainly in the embryonic liver, breast muscle, and the extraembryonic yolk sac (YS) tissue. During the last phase of incubation, as the embryo nears hatching, dynamic changes in energy metabolism occur. These changes may affect embryonic survival, hatchlings' uniformity, quality and post hatch performance of broilers, hence, being of great importance to poultry production. Here, we followed the dynamics of creatine and glycogen from embryonic day (E) 11 until hatch and up to chick placement at the farm. We showed that creatine is stored mainly in the breast muscle while glycogen is stored mainly in the YS tissue. Analysis of creatine synthesis genes revealed their expression in the liver, kidney, YS tissue and in the breast muscle, suggesting a full synthesis capacity in these tissues. Expression analysis of genes involved in gluconeogenesis, glycogenesis, and glycogenolysis, revealed that glycogen metabolism is most active in the liver. Nevertheless, due to the relatively large size of the breast muscle and YS tissue, their contribution to glycogen metabolism in embryos is valuable. Towards hatch, post E19, creatine levels in all tissues increased while glycogen levels dramatically decreased and reached low levels at hatch and at chick placement. This proves the utmost importance of creatine in energy supply to late-term embryos and hatchlings.

Intra-amniotic administration of l-glutamine promotes intestinal maturation and enteroendocrine stimulation in chick embryos.

Initial nutritional stimulation is a key driving force for small intestinal maturation. In chick embryos, administration of l-glutamine (Gln) into the amniotic fluid stimulates early development of the small intestinal epithelium by promoting enterocyte differentiation. In this study, we evaluated the effects of intra-amniotic administration of Gln on enterocyte morphology and function, and elucidated a potential enteroendocrine pathway through which Gln stimulates small intestinal maturation. Our results show that Gln stimulation at embryonic day 17 significantly increased enterocyte and microvilli dimensions by 10 and 20%, respectively, within 48 h. Post-hatch, enterocytes and microvilli were 20% longer in Gln-treated chicks. Correspondingly, Gln stimulation significantly upregulated mRNA expression of brush border nutrient transporters PepT-1 and SGLT-1 and tight junction proteins TJP-1 and TJP-2, before and after hatch (P < 0.05). Since GLP-2 signaling from intestinal L-cells is associated with enterocyte growth, functionality and integrity, we examined the effects of Gln stimulation on mRNA expression of key hormones and receptors within this enteroendocrine pathway and found significant increases in GLP-2R, IGF-1 and IGF-1R expression before and after hatch (P < 0.05). In conclusion, our findings link primary nutrient stimulation in the developing small intestine with enterocyte morphological and functional maturation and enteroendocrine signaling.

Nutritional stimulation by in-ovo feeding modulates cellular proliferation and differentiation in the small intestinal epithelium of chicks.

Nutritional stimulation of the developing small intestine of chick embryos can be conducted by in-ovo feeding (IOF). We hypothesized that IOF of glutamine and leucine can enhance small intestinal development by promoting proliferation and differentiation of multipotent small intestinal epithelial cells. Broiler embryos (n = 128) were subject to IOF of glutamine (IOF-Gln), leucine (IOF-Leu), NaCl (IOF-NaCl) or no injection (control) at embryonic d 17 (E 17). Multipotent, progenitor and differentiated cells were located and quantified in the small intestinal epithelium between E 17 and d 7 after hatch (D 7) in all treatment groups by immunofluorescence of SRY-box transcription factor 9 (Sox9) and proliferating cell nuclear antigen (PCNA), in-situ hybridization of leucine-rich repeat containing G-protein coupled receptor 5 (Lgr5) and peptide transporter 1 (PepT1) and histochemical goblet cell staining. The effects of IOF treatments at E 19 (48 h post-IOF), in comparison to control embryos, were as follows: total cell counts increased by 40%, 33% and 19%, and multipotent cell counts increased by 52%, 50% and 38%, in IOF-Gln, IOF-Leu and IOF-NaCl embryos, respectively. Only IOF-Gln embryos exhibited a significance, 36% increase in progenitor cell counts. All IOF treatments shifted Lgr5+ stem cell localizations to villus bottoms. The differentiated, PepT1+ region of the villi was 1.9 and 1.3-fold longer in IOF-Gln and IOF-Leu embryos, respectively, while goblet cell densities decreased by 20% in IOF-Gln embryos. Post-hatch, crypt and villi epithelial cell counts were significantly higher IOF-Gln chicks, compared to control chicks (P < 0.05). We conclude IOF of glutamine stimulates small intestinal maturation and functionality during the peri-hatch period by promoting multipotent cell proliferation and differentiation, resulting in enhanced compartmentalization of multipotent and differentiated cell niches and expansions of the absorptive surface area.

The transfer of amino acids and minerals to the egg yolk and to the yolk sac of their progeny is affected by breeder age.

This study aimed to assess the effects of breeder age on egg quality and amino acid and mineral transfer to the egg yolk and yolk sac of newly hatched chicks. Three ages (32, 42 and 52 weeks) of the same commercial flock of Hubbard breeders were studied. A total of 465 eggs were used for each age, with 60 being used for determining egg quality and amino acid and mineral content of yolk, and 405 for incubation period to obtain and evaluate the yolk sac of chicks. Breeders aged 52 weeks had heavier eggs and a higher percentage of yolk (p < 0.05), whereas 32-week-old breeders had higher eggshell percentage and thickness (p < 0.05). The percentage of protein deposited in egg yolk for 52-week-old breeders was higher than that for 32- and 42-week-old breeders (p < 0.05). Percentages of methionine, cysteine, met + cysteine, lysine, threonine, tryptophan, arginine and isoleucine in egg yolk for 32-week-old breeders were higher than that for 42- and 52-week-old breeders (p < 0.05). The transfer from breeder of phosphorus, potassium, calcium, magnesium, copper, iron, manganese and zinc to the yolk of eggs from 32-week-old breeders was greater than that for eggs from 42- and 52-week-old breeders (p < 0.05). Chicks from 32-week-old breeders had greater deposition of phosphorus and calcium in the yolk sac (p < 0.05). Breeder age did not affect the deposition of potassium, magnesium, copper, iron, manganese and zinc in the yolk sac of newly hatch chicks (p > 0.05). It can, however, be concluded that younger breeders deposit more amino acids and minerals in egg yolk, while embryos of older breeders seem to use the nutrients present in the yolk more efficiently during embryonic development.

Intestinal brush border assembly during the peri-hatch period and its contribution to surface area expansion.

Microvilli generate the small intestinal brush border, the main site of nutrient digestion and absorption. Mucosal structuring of the small intestine of chicken during the perihatch period has been widely researched, yet the developmental dynamics of microvilli during this period have not been fully characterized. In this study, we examined the structural and molecular characteristics of microvilli assembly and maturation during the perihatch period. Small intestines of broiler embryos and chicks were sampled at prehatch ages 17 E and 19 E, at day of hatch (DOH) and at 1, 3, 7, and 10 d posthatch. Morphological evaluations and measurements were conducted by scanning electron microscopy (SEM) and light microscopy (LM) (n = 3/timepoint), and expression of microvilli structural genes Plastin 1, Ezrin, and Myo1a was examined by Real-Time qPCR (n = 6/timepoint). Results revealed dissimilar patterns of microvilli and villi development during the perihatch period. From 19 E to 1 d, microvilli lengths increased 4.3-fold while villi lengths increased 2.8-fold (P < 0.0001). From 3 to 7 d, villi lengths increased by 20% (P < 0.005), while microvilli lengths decreased by 41% (P = 0.001). At 10 d, microvilli lengths stabilized, while villi continued to elongate by 26% (P < 0.0001). Estimations of the microvilli amplification factor (MAF) and total enterocyte surface area (TESA) revealed similar trends, with peak values of 78.53 and 1961.67 µm2, respectively, at 3 d. Microvilli structural gene expression portrayed diverse patterns. Expression of Plastin 1, which bundles and binds actin cores to the terminal web, increased 8.7-fold between 17 E and DOH (P = 0.005), and gradually increased up to 7 d (P = 0.045). Ezrin and Myo1a, both actin core-cell membrane cross-linkers, portrayed different expression patterns throughout the perihatch period, as Ezrin expression was relatively stable, while Myo1a expression increased 15.8-fold between 17 E and 10 d (P < 0.0001). We conclude that microvilli assembly during the perihatch period is a rapid, coordinated process, which dramatically expands the digestive and absorptive surface area of the small intestine before the completion of villi maturation.

Incubation temperature affects yolk utilization through changes in expression of yolk sac tissue functional genes.

The yolk sac tissue (YST) is a multifunctional metabolic organ supporting chicken embryonic development. This study examined whether incubation temperatures (ITs) affect YST functions. For this purpose, 300 eggs were assigned to 3 groups and incubated at control IT of 37.8°C, at 1.5°C below, 36.3°C (cold IT), and at 1.5°C above, 39.3°C (hot IT). For each group, 6 embryos' whole body mass and residual yolk (RSY) weights were recorded during incubation, and YST was sampled for both histology and gene expression analysis. YST functionality during incubation was examined by regression analysis, comparing changes in expression patterns of genes involved in lipid uptake and metabolism (LRP2, ApoA1), oligopeptides uptake (PepT1), gluconeogenesis (FBP1), glycogenesis (GYS2), and thyroid hormones regulation (TTR, DIO1, DIO2). Results show that hot and cold ITs affected YST gene expression and yolk utilization. PepT1 expression decreased towards hatch, in both hot and cold ITs, while in the Control IT, it reached a plateau. ApoA1 and DIO2 expression showed a moderate linear fit compared to polynomial fit in the control. GYS2 expression had no change along incubation, while in the control IT, it showed a polynomial fit. Expression of LRP2, FBP1, and DIO1 genes was affected by either cold or hot IT's. TTR expression patterns were similar in all IT groups. The variations in gene expression patterns observed in the 3 ITs can explain the changes in yolk utilization, an important parameter for hatchling quality. While the control IT showed optimal utilization, with an RSY value of 11.12% at the day of hatch, the cold and hot IT groups exhibited lower utilization with an RSY value of 18.18 and 29.99%, respectively. These findings are the first to show that ITs change the expression of key YST genes, leading to variations in yolk utilization by the embryo.

From broiler breeder hen feed to the egg and embryo: The molecular effects of guanidinoacetate supplementation on creatine transport and synthesis.

Supplementation of broiler breeder hens with beneficial additives bears great potential for affecting nutrient deposition into the fertile egg. Guanidinoacetate (GAA) is the endogenous precursor of creatine that is used as a feed additive for improving cellular energy metabolism in animal nutrition. In the present study, we have investigated whether GAA supplementation in broiler breeder feed affects creatine deposition into the hatching egg and molecular mechanisms of creatine transport and synthesis within hens and their progeny. For this, broiler breeder hens of 47 wk of age were supplemented with 0.15% GAA for 15 wk, and samples from their tissues, hatching eggs and progeny were compared with those of control, nonsupplemented hens. A significant increase in creatine content was found within the yolk and albumen of hatching eggs obtained from the GAA group, compared with the control group. The GAA group exhibited a significant increased creatine transporter gene expression compared with the control group in their small intestines and oviduct. In GAA group progeny, a significant decrease in creatine transporter expression at embryonic day 19 and day of hatch was found, compared with control group progeny. At the day of hatch, creatine synthesis genes (arginine glycine amidinotransferase and guanidinoacetate N-methyltransferase) exhibited significant decrease in expression in the GAA group progeny compared with control group progeny. These results indicate that GAA supplementation in broiler breeder feed increases its absorbance and deposition into hatching eggs, subsequently affecting GAA and creatine absorbance and synthesis within broiler progeny.

It's All About Timing: Early Feeding Promotes Intestinal Maturation by Shifting the Ratios of Specialized Epithelial Cells in Chicks.

The small intestine (SI) of chicks (Gallus gallus) matures rapidly during the initial post-hatch period and acquires digestive, absorptive, and secretive capabilities. The effects of the timing of first feeding on the quantities and distribution of specialized epithelial cells, which generate and maintain SI morphology and functionality, have not yet been examined. In this study, we identified specialized SI epithelial cell sub-types, including stem, progenitor, proliferating, and differentiated cells within crypts and villi of chicks during the first 10 days post-hatch, by in situ hybridization (ISH), immunofluorescence (IF), and histochemical staining. We then examined their quantities and ratios between day of hatch and d10 in chicks that were fed upon hatch [early feeding (EF)], compared to chicks that were fed 24 h post-hatch [delayed feeding (DF)]. Results showed that EF increased total cell quantities in the crypts and villi at days 1, 3, 7, and 10, compared to DF (p < 0.0001). At d3, EF, in comparison to DF, decreased crypt stem cell proportions (p < 0.0001), increased crypt proliferating (p < 0.01) and differentiated (p < 0.05) cell proportions, and increased villus enterocyte proportions (p < 0.01). By d10, EF increased both the quantities and proportions of villus enterocytes and goblet cells, compared to DF. We conclude that feeding upon hatch, compared to 24 h-delayed feeding, enhanced SI maturation and functionality by increasing the quantities and proportions of proliferating and differentiated cells, thus expanding the digestive, absorptive, and secretive cell populations throughout the initial post-hatch period.

From Cell to Beak: In-Vitro and In-Vivo Characterization of Chicken Bitter Taste Thresholds.

Bitter taste elicits an aversive reaction, and is believed to protect against consuming poisons. Bitter molecules are detected by the Tas2r family of G-protein-coupled receptors, with a species-dependent number of subtypes. Chickens demonstrate bitter taste sensitivity despite having only three bitter taste receptors-ggTas2r1, ggTas2r2 and ggTas2r7. This minimalistic bitter taste system in chickens was used to determine relationships between in-vitro (measured in heterologous systems) and in-vivo (behavioral) detection thresholds. ggTas2r-selective ligands, nicotine (ggTas2r1), caffeine (ggTas2r2), erythromycin and (+)-catechin (ggTas2r7), and the Tas2r-promiscuous ligand quinine (all three ggTas2rs) were studied. Ligands of the same receptor had different in-vivo:in-vitro ratios, and the ggTas2r-promiscuous ligand did not exhibit lower in-vivo:in-vitro ratios than ggTas2r-selective ligands. In-vivo thresholds were similar or up to two orders of magnitude higher than the in-vitro ones.

Detecting thresholds for bitter, umami, and sweet tastants in broiler chicken using a 2-choice test method.

The sense of taste has a key role in nutrient sensing and food intake in animals. A standardized and simple method for determination of tastant-detection thresholds is required for chemosensory research in poultry. We established a 24-h, 2-alternative, forced-choice solution-consumption method and applied it to measure detection thresholds for 3 G-protein-coupled receptor-mediated taste modalities-bitter, sweet, and umami-in chicken. Four parameters were used to determine a significant response: 1) tastant-solution consumption; 2) water (tasteless) consumption; 3) total consumption (tastant and water together); 4) ratio of tastant consumption to total consumption. Our results showed that assignment of the taste solutions and a water control to 2 bottles on random sides of the pen can be reliably used for broiler chicks, even though 47% of the chicks groups demonstrated a consistently preferred side. The detection thresholds for quinine (bitter), L-monosodium glutamate (MSG) (umami), and sucrose (sweet) were determined to be 0.3 mM, 300 mM, and 1 M, respectively. The threshold results for quinine were similar to those for humans and rodents, but the chicks were found to be less sensitive to sucrose and MSG. The described method is useful for studying detection thresholds for tastants that have the potential to affect feed and water consumption in chickens.

Bitter, sweet and umami taste receptors and downstream signaling effectors: Expression in embryonic and growing chicken gastrointestinal tract.

Taste perception is a crucial biological mechanism affecting food and water choices and consumption in the animal kingdom. Bitter taste perception is mediated by a G-protein-coupled receptor (GPCR) family-the taste 2 receptors (T2R)-and their downstream proteins, whereas sweet and umami tastes are mediated by the GPCR family -taste 1 receptors (T1R) and their downstream proteins. Taste receptors and their downstream proteins have been identified in extra-gustatory tissues in mammals, such as the lungs and gastrointestinal tract (GIT), and their GIT activation has been linked with different metabolic and endocrinic pathways in the GIT. The chicken genome contains three bitter taste receptors termed ggTas2r1, ggTas2r2, and ggTas2r7, and the sweet/umami receptors ggTas1r1 and ggTas1r3, but it lacks the sweet receptor ggTas1r2. The aim of this study was to identify and determine the expression of genes related to taste perception in the chicken GIT, both at the embryonic stage and in growing chickens. The results of this study demonstrate for the first time, using real-time PCR, expression of the chicken taste receptor genes ggTas2r1, ggTas2r2, ggTas2r7, ggTas1r1, and ggTas1r3 and of their downstream protein-encoding genes TRPM5, α-gustducin, and PLCß2 in both gustatory tissues-the palate and tongue, and extra-gustatory tissues-the proventriculus, duodenum, jejunum, ileum, cecum, and colon of embryonic day 19 (E19) and growing (21 d old) chickens. Expression of these genes suggests the involvement of taste pathways for sensing carbohydrates, amino acids and bitter compounds in the chicken GIT.

Perinatal administration of a bitter tastant influences gene expression in chicken palate and duodenum.

Bitter taste receptors (Tas2rs) and downstream effectors are responsible for mediating bitterness perception and regulation of food choice in mammals. Using RT-PCR, we demonstrated the expression of three Tas2rs and taste signal transduction molecules, α-gustducin, PLCß2, and TRPM5, in the palate, tongue, and gastrointestinal tract sections in chicken. The bitter tastant quinine activates all three chicken Tas2rs in vitro as shown using calcium-imaging assays of transfected cells. Administration of quinine postnatally or perinatally (both pre- and posthatch) to chickens increased the expression of Tas2r genes in the palate by 6.45-fold (ggTas2r1 postnatal treatment), 4.86-fold (ggTas2r1 perinatal treatment), and 4.48-fold (ggTas2r7 postnatal treatment) compared to the genes' expression in the naïve group respectively, and affected taste related gene expression in the duodenum. Whereas no-choice intake of quinine solution was not significantly lower than that of water in naïve chicks, the treatment groups postnatal, prenatal, and perinatal showed significantly lower intake of quinine by 56.1, 47.7, and 50.2%, respectively, suggesting a possible trend toward sensitization. These results open new venues toward unraveling the formative stages shaping food intake and nutrition in chicken.

Temporal transcriptome analysis of the chicken embryo yolk sac.

BACKGROUND: The yolk sac (YS) is an extra-embryonic tissue that surrounds the yolk and absorbs, digests and transports nutrients during incubation of the avian embryo as well as during early term mammalian embryonic development. Understanding YS functions and development may enhance the efficient transfer of nutrients and optimize embryo development. To identify temporal large-scale patterns of gene expression and gain insights into processes and mechanisms in the YS, we performed a transcriptome study of the YS of chick embryos on embryonic days (E) E13, E15, E17, E19, and E21 (hatch). RESULTS: 3547 genes exhibited a significantly changed expression across days. Clustering and functional annotation of these genes as well as histological sectioning of the YS revealed that we monitored two cell types: the epithelial cells and the erythropoietic cells of the YS. We observed a significant up-regulation of epithelial genes involved in lipid transport and metabolism between E13 and E19. YS epithelial cells expressed a vast array of lipoprotein receptors and fatty acid transporters. Several lysosomal genes (CTSA, PSAP, NPC2) and apolipoproteins genes (apoA1, A2, B, C3) were among the highest expressed, reflecting the intensive digestion and re-synthesis of lipoproteins in YS epithelial cells. Genes associated with cytoskeletal structure were down-regulated between E17 and E21 supporting histological evidence of a degradation of YS epithelial cells towards hatch. Expression patterns of hemoglobin synthesis genes indicated a high erythropoietic capacity of the YS between E13 and E15, which decreased towards hatch. YS histological sections confirmed these results. We also observed that YS epithelial cells expressed high levels of genes coding for plasma carrier proteins (ALB, AFP, LTF, TTR), normally produced by the liver. CONCLUSIONS: Here we expand current knowledge on developmental, nutritional and molecular processes in the YS. We demonstrate that in the final week of chick embryonic development, the YS plays different roles to support or replace the functions of several organs that have not yet reached their full functional capacity. The YS has a similar functional role as the intestine in digestion and transport of nutrients, the liver in producing plasma carrier proteins and coagulation factors, and the bone marrow in synthesis of blood cells.

Nutritional aspects of honey bee-collected pollen and constraints on colony development in the eastern Mediterranean.

Pollen is the main protein and lipid source for honey bees (Apis mellifera), and nutritionally impoverished landscapes pose a threat to colony development. To determine colony nutritional demands, we analyzed a yearly cycle of bee-collected pollen from colonies in the field and compared it to colony worker production and honey bee body composition, for the first time in social insects. We monitored monthly bee production in ten colonies at each of seven sites throughout Israel, and trapped pollen bi-monthly in five additional colonies at each of four of these sites. Pollen mixtures from each sampling date and site were analyzed for weight, total protein, total fatty acids (FAs), and FA composition. Compared to more temperate climates, the eastern Mediterranean allows a relatively high yearly colony growth of ca. 300,000-400,000 bees. Colonies at higher elevation above sea level showed lower growth rates. Queen egg-laying rate did not seem to limit growth, as peaks in capped brood areas showed that queens lay a prolific 2000 eggs a day on average, with up to 3300 eggs in individual cases. Pollen uptake varied significantly among sites and seasons, with an overall annual mean total 16.8kg per colony, containing 7.14kg protein and 677g fat. Overall mean pollen protein content was high (39.8%), and mean total FA content was 3.8%. Production cost, as expressed by the amount of nutrient used per bee, was least variable for linoleic acid and protein, suggesting these as the best descriptive variables for total number of bees produced. Linolenic acid levels in pollen during the autumn were relatively low, and supplementing colonies with this essential FA may mitigate potential nutritional deficiency. The essentiality of linoleic and linolenic acids was consistent with these FAs' tendency to be present at higher levels in collected pollen than in the expected nutrients in bee bodies, demonstrating a well-developed adjustment between pollinator nutritional demands and the nutritional value of food offered by pollinated plants.