Gap junctions in Turing-type periodic feather pattern formation.

Periodic patterning requires coordinated cell-cell interactions at the tissue level. Turing showed, using mathematical modeling, how spatial patterns could arise from the reactions of a diffusive activator-inhibitor pair in an initially homogeneous 2D field. Most activators and inhibitors studied in biological systems are proteins, and the roles of cell-cell interaction, ions, bioelectricity, etc. are only now being identified. Gap junctions (GJs) mediate direct exchanges of ions or small molecules between cells, enabling rapid long-distance communications in a cell collective. They are therefore good candidates for propagating nonprotein-based patterning signals that may act according to the Turing principles. Here, we explore the possible roles of GJs in Turing-type patterning using feather pattern formation as a model. We found 7 of the 12 investigated GJ isoforms are highly dynamically expressed in the developing chicken skin. In ovo functional perturbations of the GJ isoform, connexin 30, by siRNA and the dominant-negative mutant applied before placode development led to disrupted primary feather bud formation. Interestingly, inhibition of gap junctional intercellular communication (GJIC) in the ex vivo skin explant culture allowed the sequential emergence of new feather buds at specific spatial locations relative to the existing primary buds. The results suggest that GJIC may facilitate the propagation of long-distance inhibitory signals. Thus, inhibition of GJs may stimulate Turing-type periodic feather pattern formation during chick skin development, and the removal of GJ activity would enable the emergence of new feather buds if the local environment were competent and the threshold to form buds was reached. We further propose Turing-based computational simulations that can predict the sequential appearance of these ectopic buds. Our models demonstrate how a Turing activator-inhibitor system can continue to generate patterns in the competent morphogenetic field when the level of intercellular communication at the tissue scale is modulated.

The avian ectodermal default competence to make feathers.

Feathers originate as protofeathers before birds, in pterosaurs and basal dinosaurs. What characterizes a feather is not only its outgrowth, but its barb cells differentiation and a set of beta-corneous proteins. Reticula appear concomitantly with feathers, as small bumps on plantar skin, made only of keratins. Avian scales, with their own set of beta-corneous proteins, appear more recently than feathers on the shank, and only in some species. In the chick embryo, when feather placodes form, all the non-feather areas of the integument are already specified. Among them, midventral apterium, cornea, reticula, and scale morphogenesis appear to be driven by negative regulatory mechanisms, which modulate the inherited capacity of the avian ectoderm to form feathers. Successive dermal/epidermal interactions, initiated by the Wnt/ß-catenin pathway, and involving principally Eda/Edar, BMP, FGF20 and Shh signaling, are responsible for the formation not only of feather, but also of scale placodes and reticula, with notable differences in the level of Shh, and probably FGF20 expressions. This sequence is a dynamic and labile process, the turning point being the FGF20 expression by the placode. This epidermal signal endows its associated dermis with the memory to aggregate and to stimulate the morphogenesis that follows, involving even a re-initiation of the placode.

Transcriptome analysis of sexual dimorphism in dorsal down coloration in goslings.

BACKGROUND: In day-old Hungarian white goose goslings, there is a noticeable difference in dorsal down coloration between males and females, with females having darker dorsal plumage and males having lighter plumage. The ability to autosex day-old goslings based on their dorsal down coloration is important for managing them efficiently and planning their nutrition in the poultry industry. The aim of this study was to determine the biological and genetic factors underlying this difference in dorsal down colorationthrough histological analysis, biochemical assays, transcriptomic profiling, and q‒PCR analysis. RESULTS: Tissue analysis and biochemical assays revealed that compared with males, 17-day-old embryos and day-old goslings of female geese exhibited a greater density of melanin-containing feather follicles and a greater melanin concentration in these follicles during development. Both female and male goslings had lower melanin concentrations in their dorsal skin compared to 17-day-old embryos. Transcriptome analysis identified a set of differentially expressed genes (DEGs) (MC1R, TYR, TYRP1, DCT and MITF) associated with melanogenesis pathways that were downregulated or silenced specifically in the dorsal skin of day-old goslings compared to 17-day-old embryos, affecting melanin synthesis in feather follicles. Additionally, two key genes (MC1R and MITF) associated with feather coloration showed differences between males and females, with females having higher expression levels correlated with increased melanin synthesis and darker plumage. CONCLUSION: The expression of multiple melanogenesis genes determines melanin synthesis in goose feather follicles. The dorsal down coloration of day-old Hungarian white goose goslings shows sexual dimorphism, likely due to differences in the expression of the MC1R and MITF genes between males and females. These results could help us better understand why male and female goslings exhibit different plumage patterns.

Salivary corticosterone reflects plasmatic levels in a wild seabird.

Wild animals have been increasingly exposed to a wide range of stressors, mainly due to the intensification of human activities and habitat modifications. Consequently, new tools in order to assess the physiological and health status of wild animals have been developed. In particular, glucocorticoids have received a special attention. Primarily metabolic hormones, they are also used to evaluate the stress level of organisms. While historically measured in blood samples, new less-invasive methods have been recently developed to measure glucocorticoids in matrices such as faeces, hairs/feathers, or saliva. To date, measurements in saliva are still in their infancy despite the numerous advantages of the matrix: non-invasive, reflects the biologically active portion of glucocorticoids, allows to measure both baseline and stress-induced levels. In addition, most studies using saliva have been performed on domestic and captive animals, and recent development in wild animals have focused on mammals. Here, we show, for the first time, that saliva could also be reliably used in free-ranging birds, as glucocorticoid levels in saliva strongly correlated with plasma levels. This promising result opens new avenues for a non-invasive sampling method to assess health status of wild birds in conservation biology and ecology.

Transcriptomics Reveals the Mechanism of Purpureocillium lilacinum GZAC18-2JMP in Degrading Keratin Material.

Microbial degradation of keratin is characterized by its inherent safety, remarkable efficiency, and the production of copious degradation products. All these attributes contribute to the effective management of waste materials at high value-added and in a sustainable manner. Microbial degradation of keratin materials remains unclear, however, with variations observed in the degradation genes and pathways among different microorganisms. In this study, we sequenced the transcriptome of Purpureocillium lilacinum GZAC18-2JMP mycelia on control medium and the medium containing 1% feather powder, analyzed the differentially expressed genes, and revealed the degradation mechanism of chicken feathers by P. lilacinum GZAC18-2JMP. The results showed that the chicken feather degradation rate of P. lilacinum GZAC18-2JMP reached 64% after 216 h of incubation in the fermentation medium, reaching a peak value of 148.9 µg·mL-1 at 192 h, and the keratinase enzyme activity reached a peak value of 211 U·mL-1 at 168 h, which revealed that P. lilacinum GZAC18-2JMP had a better keratin degradation effect. A total of 1001 differentially expressed genes (DEGs) were identified from the transcriptome database, including 475 upregulated genes and 577 downregulated genes. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis of the DEGs revealed that the metabolic pathways related to keratin degradation were mainly sulfur metabolism, ABC transporters, and amino acid metabolism. Therefore, the results of this study provide an opportunity to gain further insight into keratin degradation and promote the biotransformation of feather wastes.

Biodegradation of Keratin Waste by Bacillus velezensis HFS_F2 through Optimized Keratinase Production Medium.

Enormous aggregates of keratinous wastes are produced annually by the poultry and leather industries which cause environmental degradation globally. To combat this issue, microbially synthesized extracellular proteases known as keratinase are used widely which is effective in degrading keratin found in hair and feathers. In the present work, keratinolytic bacteria were isolated from poultry farm soil and feather waste, and various cultural conditions were optimized to provide the highest enzyme production for efficient keratin waste degradation. Based on the primary and secondary screening methods, the potent keratinolytic strain (HFS_F2T) with the highest enzyme activity 32.65 ± 0.16 U/mL was genotypically characterized by 16S rRNA sequencing and was confirmed as Bacillus velezensis HFS_F2T ON556508. Through one-variable-at-a-time approach (OVAT), the keratinase production medium was optimized with sucrose (carbon source), beef extract (nitrogen source) pH-7, inoculum size (5%), and incubation at 37 °C). The degree of degradation (%DD) of keratin wastes was evaluated after 35 days of degradation in the optimized keratinase production medium devoid of feather meal under submerged fermentation conditions. Further, the deteriorated keratin wastes were visually examined and the hydrolysed bovine hair with 77.32 ± 0.32% degradation was morphologically analysed through Field Emission Scanning Electron Microscopy (FESEM) to confirm the structural disintegration of the cuticle. Therefore, the current study would be a convincing strategy for reducing the detrimental impact of pollutants from the poultry and leather industries by efficient keratin waste degradation through the production of microbial keratinase.

Effect of a novel alkaline protease from Bacillus licheniformis on growth performance, carcass characteristics, meat quality, antioxidant capacity, and intestinal morphology of white feather broilers.

BACKGROUND: The present study was conducted to investigate the effects of dietary novel alkaline protease from Bacillus licheniformis on the growth performance, meat quality, antioxidant status and intestinal morphology of broilers. In total, 4000 broilers were randomly assigned into five groups and treated with normal control, normal control + 100 mg kg-1 protease, normal control + 200 mg kg-1 protease, normal control + 300 mg kg-1 protease and normal control + 400 mg kg-1 protease. RESULTS: Supplementing protease impacted final body weight (linear, P = 0.003; quadratic, P = 0.006) and decreased feed conversion rate (linear, P = 0.036) in broilers. Moreover, dietary protease significantly increased breast muscle rate (linear, P = 0.005; quadratic, P = 0.021) and decreased drip loss (linear, P < 0.001; quadratic, P < 0.001). In addition, dietary protease notably increased protein digestibility (linear, P = 0.001; quadratic, P = 0.006) and trypsin activity (linear, P = 0.002; quadratic, P = 0.009) in jejunum. Light microscopy revealed that the jejunum villi in the 300 mg kg-1 and 400 mg kg-1 groups exhibited greater height and a denser arrangement compared to those in the control group. The addition of protease decreased malondialdehyde content (linear, P < 0.001; quadratic, P < 0.001) and increased total antioxidant capacity (linear, P = 0.001; quadratic, P < 0.001) in pectoral muscles. CONCLUSION: The results of the present study suggest that dietary novel alkaline protease from B. licheniformis improved growth performance by affecting trypsin activity, protein digestibility, antioxidant capacity and intestinal health. © 2024 Society of Chemical Industry.

Melanophilin regulates dendritogenesis in melanocytes for feather pigmentation.

Limited studies using animal models with a few natural mutations in melanophilin (Mlph) provided partial functions of Mlph in melanosome trafficking. To investigate cellular functions of Mlph, especially ZnF motif of Mlph, we analyzed all three Mlph knockout (KO) quail lines, one and two base pair (bp) deletions as models for total KO, and three bp deletion causing deletion of one Cysteine (C84del) in the ZnF motif. All quail lines had diluted feather pigmentation with impaired dendritogenesis and melanosome transport in melanocytes. In vitro studies revealed capability of binding of the ZnF motif to PIP3, and impairment of PI3P binding and mislocalization of MLPH proteins with ZnF motif mutations. The shortened melanocyte dendrites by the C84del mutation were rescued by introducing WT Mlph in vitro. These results revealed the diluted feather pigmentation by Mlph mutations resulted from congregation of melanosomes in the cell bodies with impairment of the dendritogenesis and the transport of melanosomes to the cell periphery.

Structural and enzymatic characterization of a novel metallo-serine keratinase KerJY-23.

KerJY-23 was a novel keratinase from feather-degrading Ectobacillus sp. JY-23, but its enzymatic characterization and structure are still unclear. In this study, the KerJY-23 was obtained by heterologous expression in Escherichia coli BL21(DE3), and enzymatic properties indicated that KerJY-23 was optimal at 60 °C and pH 9.0 and could be promoted by divalent metal ions or reducing agents. Furthermore, KerJY-23 had a broad substrate specificity towards casein, soluble keratin, and expanded feather powder, but its in vitro degradation against chicken feathers required an additional reducing agent. Homology modeling indicated that KerJY-23 contained a highly conserved zinc-binding HELTH motif and a His-Asp-Ser catalytic triad that belonged to the typical characteristics of M4-family metallo-keratinase and serine-keratinase, respectively. Molecular docking revealed that KerJY-23 achieved a reinforced binding on feather keratin via abundant hydrogen bonding interactions. This work not only deepened understanding of the novel and interesting metallo-serine keratinase KerJY-23, but also provided a theoretical basis for realizing the efficient use of waste feather keratin.

Testing for effects of growth rate on isotope trophic discrimination factors and evaluating the performance of Bayesian stable isotope mixing models experimentally: A moment of truth?

Discerning assimilated diets of wild animals using stable isotopes is well established where potential dietary items in food webs are isotopically distinct. With the advent of mixing models, and Bayesian extensions of such models (Bayesian Stable Isotope Mixing Models, BSIMMs), statistical techniques available for these efforts have been rapidly increasing. The accuracy with which BSIMMs quantify diet, however, depends on several factors including uncertainty in tissue discrimination factors (TDFs; Δ) and identification of appropriate error structures. Whereas performance of BSIMMs has mostly been evaluated with simulations, here we test the efficacy of BSIMMs by raising domestic broiler chicks (Gallus gallus domesticus) on four isotopically distinct diets under controlled environmental conditions, ideal for evaluating factors that affect TDFs and testing how BSIMMs allocate individual birds to diets that vary in isotopic similarity. For both liver and feather tissues, δ13C and δ 15N values differed among dietary groups. Δ13C of liver, but not feather, was negatively related to the rate at which individuals gained body mass. For Δ15N, we identified effects of dietary group, sex, and tissue type, as well as an interaction between sex and tissue type, with females having higher liver Δ15N relative to males. For both tissues, BSIMMs allocated most chicks to correct dietary groups, especially for models using combined TDFs rather than diet-specific TDFs, and those applying a multiplicative error structure. These findings provide new information on how biological processes affect TDFs and confirm that adequately accounting for variability in consumer isotopes is necessary to optimize performance of BSIMMs. Moreover, results demonstrate experimentally that these models reliably characterize consumed diets when appropriately parameterized.

Conserved regulatory switches for the transition from natal down to juvenile feather in birds.

The transition from natal downs for heat conservation to juvenile feathers for simple flight is a remarkable environmental adaptation process in avian evolution. However, the underlying epigenetic mechanism for this primary feather transition is mostly unknown. Here we conducted time-ordered gene co-expression network construction, epigenetic analysis, and functional perturbations in developing feather follicles to elucidate four downy-juvenile feather transition events. We report that extracellular matrix reorganization leads to peripheral pulp formation, which mediates epithelial-mesenchymal interactions for branching morphogenesis. α-SMA (ACTA2) compartmentalizes dermal papilla stem cells for feather renewal cycling. LEF1 works as a key hub of Wnt signaling to build rachis and converts radial downy to bilateral symmetry. Novel usage of scale keratins strengthens feather sheath with SOX14 as the epigenetic regulator. We show that this primary feather transition is largely conserved in chicken (precocial) and zebra finch (altricial) and discuss the possibility that this evolutionary adaptation process started in feathered dinosaurs.

Omics-Based Investigation on Mechanisms Controlling Cellular Internalization of Keratin Monomers during Biodegradation by Stenotrophomonas maltophilia DHHJ.

INTRODUCTION: The global poultry industry produces millions of tons of waste feathers every year, which can be bio-degraded to make feed, fertilizer, and daily chemicals. However, feather bio-degradation is a complex process that is not yet fully understood. This results in low degradation efficiency and difficulty in industrial applications. Omics-driven system biology research offers an effective solution to quickly and comprehensively understand the molecularmechanisms involved in a metabolic pathway. METHODS: In the early stage of this process, feathers are hydrolyzed into water-soluble keratin monomers. In this study, we used high-throughput RNA-seq technology to analyze the genes involved in the internalization and degradation of keratin monomers in Stenotrophomonas maltophilia DHHJ strain cells. Moreover, we used Co-IP with LC-MS/MS technology to search for proteins that interact with recombinant keratin monomers. RESULTS: We discovered TonB transports and molecular chaperones associating with the keratin monomer, which may play a crucial role in the transmembrane transport of keratin. Meanwhile, multiple proteases belonging to distinct families were identified as binding partners of keratin monomers, among which ATPases associated with diverse cellular activity (AAA+) family proteases are overrepresented. Four genes, including JJL50_15620, JJL50_17955 (TonB-dependent receptors), JJL50_03260 (ABC transporter ATP-binding protein), and JJL50_20035 (ABC transporter substrate-binding protein), were selected as representatives for determining their expressions under different culture conditions using qRT-PCR, and they were found to be upregulated in response to keratin degradation consistent with the data from RNA-seq and Co-IP. CONCLUSION: This study highlights the complexity of keratin biodegradation in S. maltophilia DHHJ, in which multiple pathways are involved such as protein folding, protein transport, and several protease systems. Our findings provide new insights into the mechanism of feather degradation.

Effects of Feather Hydrolysates Generated by Probiotic Bacillus licheniformis WHU on Gut Microbiota of Broiler and Common carp.

Due to the ever-increasing demand for meat, it has become necessary to identify cheap and sustainable sources of protein for animal feed. Feathers are the major byproduct of poultry industry, which are rich in hard-to-degrade keratin protein. Previously we found that intact feathers can be digested into free amino acids, short peptides, and nano-/micro-keratin particles by the strain Bacillus licheniformis WHU in water, and the resulting feather hydrolysates exhibit prebiotic effects on mice. To explore the potential utilization of feather hydrolysate in the feed industry, we investigated its effects on the gut microbiota of broilers and fish. Our results suggest that feather hydrolysates significantly decrease and increase the diversity of gut microbial communities in broilers and fish, respectively. The composition of the gut microbiota was markedly altered in both of the animals. The abundance of bacteria with potentially pathogenic phenotypes in the gut microbial community of the fish significantly decreased. Staphylococcus spp., Pseudomonas spp., Neisseria spp., Achromobacter spp. were significantly inhibited by the feather hydrolysates. In addition, feather hydrolysates significantly improved proteolytic activity in the guts of broilers and fish. In fish, the expression levels of ZO-1 and TGF-α significantly improved after administration of feather hydrolysates. The results presented here suggest that feather hydrolysates generated by B. licheniformis WHU could be an alternative protein source in aquaculture and could exert beneficial effects on fish.