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Objective: Transgenic hens hold a great promise to produce various valuable proteins. Through virus transduction into stage X embryo, the transgene expression under the control of constructed chicken ovalbumin promoters has been successfully achieved. However, a validation system that can evaluate differently developed ovalbumin promoters in in vitro, remains to be developed. Methods: In the present study, chicken oviduct epithelial cells (cOECs) were isolated from oviduct tissue and shortly cultured with keratinocyte complete medium supplemented with chicken serum. The isolated cells were characterized with immunofluorescence, western blot, and flow cytometry using oviduct-specific marker. Chicken mutated ovalbumin promoter (Mut-4.4-kb-pOV) was validated in these cells using luciferase reporter analysis. Results: The isolated cOECs revealed that the oviduct-specific marker, ovalbumin protein, was clearly detected by immunofluorescence, western blot, and flow cytometry analysis revealed that approximately 79.40% of the cells contained this protein. Also, luciferase reporter analysis showed that the constructed Mut-4.4-kb-pOV exhibited 7.1-fold (pin vitro validation of the recombinant promoter activity in cOECs can facilitate the production of efficient transgenic chickens for potential use as bioreactors.

Campylobacter jejuni is the leading cause of bacterial gastroenteritis globally, and infections are often transmitted through consumption of raw or undercooked poultry. Campylobacter jejuni ST50 is among the top ten sequence types (STs) reported in the collected isolates listed at PubMLST records from poultry, food and clinical sources for Asia, Europe, North America, Oceania and South America. This study was designed to determine the most commonly reported C. jejuni STs globally using the PubMLST database and assess similarities between genomes of C. jejuni ST50 isolates from geographically distinct locations. To gain a better understanding of C. jejuni diversity, we compared draft genome sequences of 182 ST50 isolates recovered from retail or caecal poultry samples in Oceania, Europe and North America that were collected over a period of 9 years (2010 to 2018). Overall, phylogenetic analysis revealed that isolates from geographically distinct locations tended to cluster based on the continent where the sample was collected. Among ST50 isolates from Europe and North America, we identified resistance determinants associated with phenotypic resistance to beta-lactams (EU: 55%; GB: 43.1%), tetracyclines (CA: 77.3%; EU: 37.5%; GB: 9.8%; US: 43.5%) and fluoroquinolones (EU: 60.0%; GB: 15.7%); no resistance determinants were identified in isolates from Australia. In general, the majority of the virulence genes, with rare exceptions such as wlaN, cj1138, hddA and rfbC, were evenly distributed throughout the genomes of all ST50 isolates in this study. Genomic-based characterization of C. jejuni ST50 isolates from poultry on three continents highlighted that geographically distinct isolates have evolved independently but only represent a glimpse into the diversity of C. jejuni.

Antimicrobials are extensively used both prophylactically and therapeutically in poultry production. Despite this, there are little data on the effect of antimicrobial use (AMU) on disease incidence rate and per cent mortality. We investigated the relationships between AMU and disease and between AMU and mortality using data from a large (n = 322 flocks) cohort of small-scale chicken flocks in the Mekong Delta, Vietnam, that were followed longitudinally from day old to slaughter (5,566 observation weeks). We developed a parameterized algorithm to emulate a randomized control trial from observational data by categorizing the observation weeks into 'non-AMU', 'prophylactic AMU' and 'therapeutic AMU'. To evaluate the prophylactic AMU effect, we compared the frequencies of clinical signs in 'non-AMU' and 'prophylactic AMU' periods. To analyse therapeutic AMU, we compared weekly per cent mortality between the weeks of disease episodes before and after AMU. Analyses were stratified by clinical signs (4) and antimicrobial classes (13). Prophylactic AMU never reduced the probability of disease, and some antimicrobial classes such as lincosamides, amphenicols and penicillins increased the risk. The risk of diarrhoea consistently increased with prophylactic AMU. Therapeutic AMU often had an effect on mortality, but the pattern was inconsistent across the combinations of antimicrobial classes and clinical signs with 14/29 decreasing and 11/29 increasing the per cent weekly mortality. Lincosamides, methenamines and cephalosporins were the only three antimicrobial classes that always decreased the mortality when used therapeutically. Results were robust respective to the parameters values of the weeks categorization algorithm. This information should help support policy efforts and interventions aiming at reducing AMU in animal production.

Background A body distribution with high intramuscular fat and low abdominal fat is the ideal goal for broiler breeding. Preadipocytes with different origins have differences in terms of metabolism and gene expression. The transcriptome analysis performed in this study of intramuscular preadipocytes (DIMFPs) and adipose tissue-derived preadipocytes (DAFPs) aimed to explore the characteristics of lipid deposition in different chicken preadipocytes by dedifferentiation in vitro. Results Compared with DAFPs, the total lipid content in DIMFPs was reduced (P ). Moreover, 72 DEGs related to lipid metabolism were screened, which were involved in adipocyte differentiation, fatty acid transport and fatty acid synthesis, lipid stabilization, and lipolysis. Among the 72 DEGs, 19 DEGs were enriched in the PPAR signaling pathway, indicating its main contribution to the regulation of the difference in lipid deposition between DAFPs and DIMFPs. Among these 19 genes, the representative APOA1, ADIPOQ, FABP3, FABP4, FABP7, HMGCS2, LPL and RXRG genes were downregulated, but the ACSL1, FABP5, PCK2, PDPK1, PPARG, SCD, SCD5, and SLC27A6 genes were upregulated (P P Conclusions Our data suggested a difference in lipid deposition between DIMFPs and DAFPs of chickens in vitro and proposed a molecular regulatory network for the difference in lipid deposition between chicken DAFPs and DIMFPs. The lipid content was significantly increased in DAFPs by the direct mediation of PPAR signaling pathways. These findings provide new insights into the regulation of tissue-specific fat deposition and the optimization of body fat distribution in broilers.

Objective: Glucose transporter 9 (GLUT9) is a uric acid transporter that is associated with uric absorption in mice and humans; but it is unknown whether GLUT9 involves in chicken uric acid regulation. This experiment aimed to investigate the chicken GLUT9 expression and serum uric acid (SUA) level.Methods: Sixty chickens were divided into 4 groups (n = 15): a control group (NC); a sulfonamide-treated group (SD) supplemented with sulfamonomethoxine sodium via drinking water (8 mg/L); a fishmeal group (FM) supplemented with 16% fishmeal in diet; and a uric acid-injection group (IU), where uric acid (250 mg/kg) was intraperitoneally injected once a day. The serum was collected weekly to detect the SUA level. Liver, kidney, jejunum, and ileum tissues were collected to detect the GLUT9 mRNA and protein expression.Results: The results showed in the SD and IU groups, the SUA level increased and GLUT9 expression increased in the liver, but decreased in the kidney, jejunum, and ileum. In the FM group, the SUA level decreased slightly and GLUT9 expression increased in the kidney, but decreased in the liver, jejunum, and ileum. Correlation analysis revealed that liver GLUT9 expression correlated positively, and renal GLUT9 expression correlated negatively with the SUA level.Conclusion: These results demonstrate that there may be a feedback regulation of GLUT9 in the chicken liver and kidney to maintain the SUA balance; however, the underlying mechanism needs to be investigated in future studies.

Avian species are used as model systems in research and have contributed to ground-breaking concepts in developmental biology, immunology, genetics, virology, cancer and cell biology. The chicken in particular is an important research model and an agricultural animal as a major contributor to animal protein resources for the global population. The development of genome editing methods, including CRISPR/Cas9, to mediate germline engineering of the avian genome will have important applications in biomedical, agricultural and biotechnological activities. Notably, these precise genome editing tools have the potential to enhance avian health and productivity by identifying and validating beneficial genetic variants in bird populations. Here, we present a concise description of the existing methods and current applications of the genome editing tools in bird species, focused on chickens, with attention on animal use and welfare issues for each of the techniques presented.

BackgroundInappropriate use of antimicrobial agents in animal production has led to the development of antimicrobial resistance (AMR) in foodborne pathogens. Transmission of AMR foodborne pathogens from reservoirs, particularly chickens to the human population does occur. Recently, we reported that occupational exposure was a risk factor for multidrug-resistant (MDR)Escherichia coli (E. coli)among poultry-workers. Here we determined the prevalence and genetic relatedness among MDRE. coliisolated from poultry-workers, chickens, and poultry environments in Abuja, Nigeria. This study was conducted to address the gaps identified by the Nigerian AMR situation analysis.MethodsWe conducted a cross-sectional study among poultry-workers, chickens, and poultry farm/live bird market (LBM) environments. The isolates were tested phenotypically for their antimicrobial susceptibility profiles, genotypically characterized using whole-genome sequencing (WGS) and in silico multilocus sequence types (MLST). We conducted a phylogenetic single nucleotide polymorphism (SNPs) analysis to determine relatedness and clonality among the isolates.ResultsA total of 115 (26.8%) out of 429 samples were positive forE. coli.Of these, 110 isolates were viable for phenotypic and genotypic characterization. The selection comprised 47 (42.7%) isolates from poultry-workers, 36 (32.7%) from chickens, and 27 (24.5%) from poultry-farm or LBM environments. Overall, 101 (91.8%) of the isolates were MDR conferring resistance to at least three drug classes.High frequency of resistance was observed for tetracycline (n = 102; 92.7%), trimethoprim/sulfamethoxazole (n = 93; 84.5%), streptomycin (n = 87; 79.1%) and ampicillin (n = 88; 80%). Two plasmid-mediated colistin genes—mcr-1.1harboured on IncX4 plasmids were detected in environmental isolates. The most prevalent sequence types (ST) were ST-155 (n = 8), ST-48 (n = 8) and ST-10 (n = 6). Two isolates of human and environmental sources with a SNPs difference of 6161 originating from the same farm shared a novel ST. The isolates had similar AMR genes and plasmid replicons.ConclusionMDRE.coliisolates were prevalent amongst poultry-workers, poultry, and the poultry farm/LBM environment. The emergence of MDRE. coliwith novel ST in two isolates may be plasmid-mediated. Competent authorities should enforce AMR regulations to ensure prudent use of antimicrobials to limit the risk of transmission along the food chain.

The study aimed to monitor parasite and host gene expression during the early stages of Eimeria tenella infection of chicken cells using dual RNA-Seq analysis. For this, we used chicken macrophage-like cell line HD11 cultures infected in vitro with purified E. tenella sporozoites. Cultures were harvested between 2 and 72 h post-infection and mRNA was extracted and sequenced. Dual RNA-Seq analysis showed clear patterns of altered expression for both parasite and host genes during infection. For example, genes in the chicken immune system showed upregulation early (2–4 h), a strong downregulation of genes across the immune system at 24 h and a repetition of early patterns at 72 h, indicating that invasion by a second generation of parasites was occurring. The observed downregulation may be due to immune self-regulation or to immune evasive mechanisms exerted by E. tenella. Results also suggested pathogen recognition receptors involved in E. tenella innate recognition, MRC2, TLR15 and NLRC5 and showed distinct chemokine and cytokine induction patterns. Moreover, the expression of several functional categories of Eimeria genes, such as rhoptry kinase genes and microneme genes, were also examined, showing distinctive differences which were expressed in sporozoites and merozoites.

Intestinal mucosa is the largest immune organ in animals, and its immune function is directly related to the resistance against various diseases. Taishan Pinus massoniana pollen polysaccharides (TPPPS) have been recognized as an effective vaccine adjuvant and potential immune enhancer against viral infections. However, little is known about their direct immune-enhancing activity on intestinal mucosa. In this study, we extracted the polysaccharides from Taishan masson pine pollen to investigate its promotive effect on intestinal mucosal immunity. A total of 120 1-day-old chickens were divided into 4 groups and inoculated with PBS or 3 different doses of TPPPS (10 mg/mL, 20 mg/mL, and 40 mg/mL), respectively. Feces, intestinal specimens, and serum samples were collected from the chickens at 7, 14, and 21 d after inoculation. The antibodies in serum, mucosal secretion of IgA, structure of intestinal villi, and expressions of cytokine genes and mucosal immune-related genes in the chickens were all significantly improved by TPPPS treatments. At 21 d after inoculation following the challenge of Newcastle disease virus, the chickens inoculated with 20 and 40 mg/mL TPPPS exhibited decreased weight loss and reduced intestinal pathologic damage and viral loads in the intestine. In summary, our results demonstrate that TPPPS can enhance mucosal immunity and promote intestinal villi development. This study has established the foundation for the development of novel immune-enhancing agent with immune-regulatory effects on intestinal mucosa.

Objective: Vanin1 (VNN1) is a pantetheinase that can catalyze the hydrolysis of pantetheine to produce pantothenic acid and cysteamine. Our previous studies showed that VNN1 is specifically expressed in chicken liver. In this study, we aimed to investigate the roles of peroxisome proliferators activated receptor α (PPARα) and miRNA-181a-5p in regulating VNN1 gene expression in chicken liver.Methods: 5′-RACE was performed to identify the transcription start site of chicken VNN1. JASPAR and TFSEARCH were used to analyze the potential transcription factor binding sites in the promoter region of chicken VNN1 and miRanda was used to search miRNA binding sites in 3′ untranslated region (3′UTR) of chicken VNN1. We used a knock-down strategy to manipulate PPARα (or miRNA-181a-5p) expression levels in vitro to further investigate its effect on VNN1 gene transcription. Luciferase reporter assays were used to explore the specific regions of VNN1 targeted by PPARα and miRNA-181a-5p.Results: Sequence analysis of the VNN1 promoter region revealed several transcription factor-binding sites, including hepatocyte nuclear factor 1α (HNF1α), PPARα, and CCAAT/enhancer binding protein α. GW7647 (a specific agonist of PPARα) increased the expression level of VNN1 mRNA in chicken primary hepatocytes, whereas knockdown of PPARα with siRNA increased VNN1 mRNA expression. Moreover, the predicted PPARα-binding site was confirmed to be necessary for PPARα regulation of VNN1 gene expression. In addition, the VNN1 3′UTR contains a sequence that is completely complementary to nucleotides 1 to 7 of miRNA-181a-5p. Overexpression of miR-181a-5p significantly decreased the expression level of VNN1 mRNA.Conclusion: This study demonstrates that PPARα is an important transcriptional activator of VNN1 gene expression and that miRNA-181a-5p acts as a negative regulator of VNN1 expression in chicken hepatocytes.

Do faster slaughter line speeds for young chickens increase risk of Salmonella contamination? We analyze data collected in 2018–2019 from 97 slaughter establishments processing young chickens to examine the extent to which differences in slaughter line speeds across establishments operating under the same inspection system explain observed differences in their microbial quality, specifically frequencies of positive Salmonella samples. A variety of off-the-shelf statistical and machine learning techniques applied to the data to identify and visualize correlations and potential causal relationships among variables showed that the presence of Salmonella or other indicators of process control, such as noncompliance records for regulations associated with process control and food safety, are not significantly increased in establishments with higher line speeds (e.g., above 140 birds per min) compared with establishments with lower line speeds when establishments are operating under the conditions present in this study. This included some establishments operating under specific criteria to obtain a waiver for line speed. A null hypothesis advanced over 30 yr ago by the National Research Council that increased line speeds result in a product that is not contaminated more often than before line speeds were increased, appears to be fully consistent with these recent data.

The avian eggshell is a natural protective envelope that relies on the phenomenon of biomineralization for its formation. The shell is made of calcium carbonate in the form of calcite, which contains hundreds of proteins that interact with the mineral phase controlling its formation and structural organization, and thus determine the mechanical properties of the mature biomaterial. We describe its mineralogy, structure and the regulatory interactions that integrate the mineral and organic constituents. We underline recent evidence for vesicular transfer of amorphous calcium carbonate (ACC), as a new pathway to ensure the active and continuous supply of the ions necessary for shell mineralization. Currently more than 900 proteins and thousands of upregulated transcripts have been identified during chicken eggshell formation. Bioinformatic predictions address their functionality during the biomineralization process. In addition, we describe matrix protein quantification to understand their role during the key spatially- and temporally- regulated events of shell mineralization. Finally, we propose an updated scheme with a global scenario encompassing the mechanisms of avian eggshell mineralization. With this large dataset at hand, it should now be possible to determine specific motifs, domains or proteins and peptide sequences that perform a critical function during avian eggshell biomineralization. The integration of this insight with genomic data (nonsynonymous single nucleotide polymorphisms) and precise phenotyping (shell biomechanical parameters) on pure selected lines will lead to consistently better-quality eggshell characteristics for improved food safety. This information will also address the question of how the evolutionary-optimized chicken eggshell matrix proteins affect and regulate calcium carbonate mineralization as a good example of biomimetic and bio-inspired material design., French National Research Agency (ANR) European Commission ANR-13-BSV-0007-01, Natural Sciences and Engineering Research Council of Canada (NSERC) RGPIN-2016-04410, Le STUDIUM, Centre Val de Loire, Nouzilly, France - University of Tours

The aim of the study was to understand the dynamic changes in daily step counts (DSC) during the development of chickens and to further explore the effects of exercise on the growth performance, carcass yield, meat quality, and tibial strength of cocks. A total of 600 (half male and half female) 1-day-old Wannan chickens with similar hatching weights were raised under the same rearing conditions. All birds were wing banded and housed in identical cages for from 1 to 8 wk in the experimental poultry house. The dimensions of the cages were 70 × 70 × 40 cm (length × width × height). At the age of 9 to 16 wk, these birds were reared in indoor pens (2 m × 2 m, 1,000 cm2 per bird). In addition, they also had a free-range grass paddock (20 m × 30 m, 1 m2 per bird). The DSC of male and female Wannan chicks were recorded from 70 to 112 d by using a pedometer. At 112 d of age, based on the average DSC, birds were divided into groups representing the highest (HS), medium, and lowest (LS) number of step groups. Fifteen cocks from each group were selected for subsequent experiments. Compared with the LS group, the HS group displayed higher tibial strength (P = 0.025) and lower BW, cooking loss (P = 0.014), shear force (P = 0.023), and drip loss (P = 0.008). The DSC had no effects on the female BW or male carcass parameters. There was no significant change in the DSC of all birds from 70 to 112 d. However, male chickens took more steps than females at 15 (P = 0.025) and 16 (P = 0.012) week of age. In conclusion, the effects of the DSC on the BW of Wannan chickens depend on sex, and enhanced exercise could improve the meat quality and tibial strength of cocks.

The aim of the present study was to evaluate the effects of quantitative feed restriction (FR) in fast-, medium-, and slow-growing meat-type male and female chickens on their growth, feed consumption, economic efficiency, carcass composition, and gastrointestinal microbiota. In the experiment, fast-growing Ross 308, medium-growing Hubbard JA 757 and slow-growing ISA Dual chickens of both sexes were exposed to quantitative FR between 14 and 21 d of age. During the FR, restricted chickens received 70% of the amount of feed consumed by the ad libitum (AL) group. Live weight at the end of the experiment was affected by genotype (P

Salmonella enterica serovar Gallinarum biovar Gallinarum (S. Gallinarum) is a host-specific pathogen causing systemic infection in poultry, which leads to significant economic losses due to high mortality. However, little is known about the dynamic process of systemic infection and pathogenic characteristics of S. Gallinarum in chickens. In the present study, we developed an oral infection model that reproduces the pathology of S. Gallinarum and clarified the host immune response of the infected chickens. Chickens at 20 days of age orally inoculated at a dose of 108 colony forming unit (CFU) showed typical clinical signs of fowl typhoid and died between 6 and 10 days post infection. The inoculated S. Gallinarum rapidly disseminated to multple organs and the bacterial counts increased in the liver and spleen at 3 days post infection. Pathological changes associated wirh inflammation in the liver and spleen became apparent at 4 days post infection, and increased expression of interferon (IFN)-γ and interleuikin (IL)-12 in the liver and spleen did not observed until 3 days post infection. These results indicate that S. Gallinarum rapidly spread to entire body through intestine, and the low-level of inflammatory responses in the liver during the early stage of infection may contribute to rapid, systemic dissemination of the bacteria. Our infection model and findings will contribute to the better understanding of the pathogenic mechanism of S. Gallinarum, and provide new insights into the prevention and control of fowl typhoid.

Background DNA methylation is a significant epigenetic modification that is evolutionarily conserved in various species and often serves as a repressive mark for transcription. DNA methylation levels and patterns are regulated by a balance of opposing enzyme functions, DNA methyltransferases, DNMT1/3A/3B and methylcytosine dioxygenases, TET1/2/3. In mice, the TET enzyme converts DNA cytosine methylation (5mC) to 5-hydroxymethylcytosine (5hmC) at the beginning of fertilisation and gastrulation and initiates a global loss of 5mC, while the 5mC level is increased on the onset of cell differentiation during early embryonic development. Objective Global loss and gain of DNA methylation may be differently regulated in diverged species. Methods Chicken B-cell lymphoma DT40 cells were used as an avian model to compare differences in the overall regulation of DNA modification with mammals. Results We found that DNA methylation is maintained at high levels in DT40 cells through compact chromatin formation, which inhibits TET-mediated demethylation. Human and mouse chromosomes introduced into DT40 cells by cell fusion lost the majority of 5mC, except for human subtelomeric repeats. Conclusion Our attempt to elucidate the differences in the epigenetic regulatory mechanisms between birds and mammals explored the evidence that they share a common chromatin-based regulation of TET–DNA access, while chicken DNMT1 is involved in different target sequence recognition systems, suggesting that factors inducing DNMT–DNA association have already diverged.

Graphical abstract THQ of Cr in the meat was more than one. HI in chicken was more than one. CR was neglectable in the case of both Cd and Pb., Highlights • All of the samples were contaminated with Cd, Cr, Cu, Ni and Pb. • The highest concentration of Pb (0.11 mg/kg) was in the gizzard. • The concentration of Cd and Cu were highest in the liver, while the highest contents of Cr and Ni were in the chicken. • THQs of Cr in the muscle were more than one. • Consumption of chicken meat in both of adults and children may pose a significant health risk for consumers., The present study aimed to determine the average concentration of some metals, including cadmium (Cd), chromium (Cr), copper (Cu), nickel (Ni) and lead (Pb) in the chicken, hen’s liver, and gizzard in the east of Iran. Estimated daily intake (EDI), target hazard quotient (THQ), hazard index (HI) and carcinogenic risk (CR) were calculated. In this cross-sectional study, fifty one samples including chicken, hen’s liver and gizzard were obtained from Birjand, Iran. Measurement of Cd, Cr, Cu, Ni, and Pb was carried out by using an Inductively Coupled Plasma-Optic Emission Spectroscopy (ICP-OES). All of the measured metals were detected in 100 % of the samples. The metals had a different distribution pattern. The highest concentration of Cd and Cu was in the liver samples while the Cr and Ni had the highest levels in the chicken. Pb concentration was at the highest level in the gizzard. The least amount of Cr, Ni, and Pb was found in the liver while Cu had the least content in the muscle. EDI had an acceptable level, but the highest daily intake of all studied metals was through muscle. Cr had the highest THQ and it was more than one in the meat. HI in chicken was more than one. Liver and gizzard of hens had a neglectable HI. CR was neglectable in the case of both Cd and Pb, but it was considerable for Cr and Ni. The consumption of chicken in both adults and children may pose a significant health risk for consumers.

Objective: To explore the molecular mechanisms of fatty liver hemorrhagic syndrome (FLHS) in laying hens, an experiment was conducted to reveal the differences in histopathological observation and gene expression between FLHS group and normal group.Methods: We compared the histopathological difference using hematoxylin and eosin staining and proceeded with RNA sequencing of adipose tissue to search differentially expressed genes and enriched biological processes and pathways. Then we validated the mRNA expression levels by real-time polymerase chain reaction and quantified protein levels in the circulation by enzyme-linked immunosorbent assay.Results: We identified 100 differentially expressed transcripts corresponding to 66 genes (DEGs) were identified between FLHS-affected group and normal group. Seven DEGs were significantly enriched in the immune response process and lipid metabolic process, including phospholipase A2 group V, WAP kunitz and netrin domain containing 2, delta 4-desaturase sphingolipid 2, perilipin 3, interleukin-6 (IL-6), ciliary neurotrophic factor (CNTF), and suppressor of cytokine signaling 3 (SOCS3). And these genes could be the targets of immune response and be involved in metabolic homeostasis during the process of FLHS in laying hens. Based on functional categories of the DEGs, we further proposed a model to explain the etiology and pathogenesis of FLHS. IL-6 and SOCS3 mediate inflammatory responses and the satiety hormone of leptin, induce dysfunction of Jak-STAT signaling pathway, leading to insulin resistance and lipid metabolic disorders. Conversely, CNTF may reduce tissue destruction during inflammatory attacks and confer protection from inflammation-induced insulin resistance in FLHS chickens.Conclusion: These findings highlight the therapeutic implications of targeting the JAK-STAT pathway. Inhibition of IL6 and SOCS3 and facilitation of CNTF could serve as a favorable strategy to enhance insulin action and improve glucose homoeostasis, which are of importance for treating obesity-related disorders for chickens.

Oral processing behaviour can affect the bioavailability of macronutrients. The aim of the present study was to determine the influence of oral processing behaviour on bolus properties and in vitro protein digestion of chicken and soya-based vegetarian chicken. Natural chewing time and chewing frequency of both foods were determined in healthy adults (n 96). While natural chewing time differed considerably between consumers (chicken: 7·7–39·4 s; soya-based vegetarian chicken: 7·8–46·2 s), chewing frequency (1·4 chews/s) did not differ considerably between consumers and was independent of product type. Natural chewing times of 11 and 24 s were found for clusters of consumers showing shortest and longest chewing time for both products. Chicken and soya-based vegetarian chicken were chewed for 11 and 24 s and boli expectorated by n 16 consumers to determine in vitro gastric digestion and by n 7 to determine in vitro intestinal digestion. For both foods, longer chewing time resulted in the formation of significantly (P in vitro degree of protein hydrolysis (DH%) than shorter chewing time (chicken: DH%11s = 7 ± 23 % and DH%24s = 89 ± 26 %; soya-based vegetarian chicken: DH%11s = 57 ± 18 % and DH%24s = 70 ± 21 %, P In vitro degree of protein hydrolysis was higher for chicken than that for soya-based vegetarian chicken regardless of chewing time. We conclude that naturally occurring longer chewing time leads to more and smaller bolus particles of chicken and soya-based vegetarian chicken and thereby increases in vitro protein hydrolysis compared with shorter chewing time.

The immunomodulatory effect of Acanthopanax senticosus polysaccharide (ASPS) on immunosuppressed chickens induced by cyclophosphamide (Cy) was observed in this study. Four hundred 7-day-old chickens were randomly divided into 4 groups: vaccinated control group (VC group), Cy-challenged control group (Cy group), Cy-challenged + low-dose ASPS group (ASPSL + Cy group), and Cy-challenged + high-dose ASPS group (ASPSH + Cy group). All groups except the VC group were injected with Cy at a dose of 80 mg/kg/day of BW for 3 successive days to induce immunosuppression. At the age of 10 d, the ASPSL + Cy group and ASPSH + Cy group were intramuscularly injected with 0.2 mL of ASPS at the dose of 100 and 200 mg/mL/day, respectively, once a day for 3 successive days. The Cy group was injected with saline solution in the same way as the 2 ASPS groups. At the age of 14 d, the chickens were vaccinated with Newcastle disease (ND) vaccine in all groups. On day 7, 14, 21, and 28 after the vaccination, BW, lymphocyte proliferation, the serum antibody titers of the ND vaccine, the proportion of CD4+ and CD8+ T lymphocytes, and the concentrations of interferon gamma and IL-2 were determined. The results showed that chickens were injected with Cy at a dose of 80 mg/kg of BW for 3 d displayed lower immune responses than the control group, indicating that the immunosuppressive model was successfully established. At most time points, both high and low doses of ASPS could significantly promote lymphocyte proliferation; enhance BW, antibody titers, and the proportion of CD4+ and CD8+ T lymphocytes; and raised the concentrations of interferon gamma and IL-2 in Cy-treated chickens compared with those in the Cy control group (P < 0.05). These results indicated that ASPS could resist immunosuppression induced by Cy and may be a new-type immune adjuvant to improve vaccination in normal and immunosuppressed chickens.

Two categories of immune responses—innate and adaptive immunity—have both polygenic backgrounds and a significant environmental component. The goal of the reported study was to define candidate genes and mutations for the immune traits of interest in chickens using machine learning–based sensitivity analysis for single-nucleotide polymorphisms (SNPs) located in candidate genes defined in quantitative trait loci regions. Here the adaptive immunity is represented by the specific antibody response toward keyhole limpet hemocyanin (KLH), whereas the innate immunity was represented by natural antibodies toward lipopolysaccharide (LPS) and lipoteichoic acid (LTA). The analysis consisted of 3 basic steps: an identification of candidate SNPs via feature selection, an optimisation of the feature set using recursive feature elimination, and finally a gene-level sensitivity analysis for final selection of models. The predictive model based on 5 genes (MAPK8IP3 CRLF3, UNC13D, ILR9, and PRCKB) explains 14.9% of variance for KLH adaptive response. The models obtained for LTA and LPS use more genes and have lower predictive power, explaining respectively 7.8 and 4.5% of total variance. In comparison, the linear models built on genes identified by a standard statistical analysis explain 1.5, 0.5, and 0.3% of variance for KLH, LTA, and LPS response, respectively. The present study shows that machine learning methods applied to systems with a complex interaction network can discover phenotype-genotype associations with much higher sensitivity than traditional statistical models. It adds contribution to evidence suggesting a role of MAPK8IP3 in the adaptive immune response. It also indicates that CRLF3 is involved in this process as well. Both findings need additional verification.

We conducted a cytological search for amyloid structures in female reproductive cells of Gallus gallus domesticus and Drosophila melanogaster. We have shown that the amyloid-specific dye, Thioflavin S, but not Congo red, stains some cytoplasmic and even nuclear structures in chicken ovaries. In fruit fly eggs both Thioflavin S and Congo red specifically stain eggshell structures such as micropyle, dorsal appendages and pillars. Moreover, these structures, when stained with Congo red, demonstrate birefringence in polarized light, which is a characteristic feature of all classical amyloids. Our data show that female reproductive cells during evolution began to use amyloid fibrils to form various functional structures necessary for development under certain environmental conditions.

The prevalence of crossed beaks ranging from 0.2 to 7.4% was documented in at least 12 chicken strains. Previous studies focused largely on candidate molecules, whereas the morphological observation was missing. This study reported a detailed phenotype and prevalence of crossed beaks based on morphological observation in nine thousand nine hundred 1-day-old female Beijing-You chicks. Affected chicks were classified into 2 categories based on the direction of the mandibular deformation: left and right. Each category was selected to sacrifice for the measurement of length, width, and thickness of the bilateral mandibular ramus (MR). The normal chicks were used as controls. Paraffin section was made for the bilateral MR of a crossed beak and a normal control for histology analysis. A total of 97 out of 9,900 chickens showed beak deformity including 71 crossed beaks (0.72%) and 26 side beaks (0.26%) for which the upper and lower beak were both bent in the same direction. There was no difference in the direction of the bend of the lower beak in crossed beaks (P > 0.05). The incidence of crossed beaks increased quickly from 0 to 56 d and no new incidence after 56 d. The angle of the crossed beaks was below 5° in the first week and had grown more severe with age until 56 d. The mandible structure showed that condyle served as a growth center for the MR extension. The short-side MR of crossed beaks was thicker than normal ones (P

The removal of antibiotics from the feeds used in the livestock industry has resulted in the use of a wide range of alternative antimicrobial products that aim to deliver the productivity and health benefits that have traditionally been associated with antibiotics. Amongst the most popular alternatives are phytogenic product-based extracts from herbs and spices with known antimicrobial properties. Despite embracing such alternatives, the industry is still largely unaware of modes of action, their overall effects on animal health, and interactions with other feed additives such as probiotics. To address some of these issues, three phytogenic products were selected and their interactions with caecal microbiota of layers, grown under six different production systems, were investigated in vitro. Caecal microbiotas were grown with and without phytogenic products, and the changes in microbiota composition were monitored by sequencing of 16S rRNA gene amplicons. Phytogenic products and production system both significantly influenced microbiota composition. The three phytogenic products all altered the relative abundance of species within the Lactobacillus genus, by promoting the growth of some and inhibiting other Lactobacillus species. There were also significant alterations in the Bacillus genus. This was further investigated by comparing the effects of the phytogenic products on the growth of a commercially used Bacillus-based probiotic. The phytogens affected the probiotic mix differently, with some promoting the growth of Bacillus sp. at lower phytogenic concentrations, and fully suppressing growth at higher concentrations, indicating the importance of finding an optimal concentration that can control pathogens while promoting beneficial bacteria. Key points • After removal of antibiotics from animal feed, urgent solutions for pathogen control were needed. • Alternative products entered the market without much knowledge on their effects on animal health. • Probiotic products are used in combination with phytogens despite the possible incompatibility.

To investigate the effects of dietary taurine supplementation on growth performance, antioxidant status, and lipid metabolism in broilers, 384 male broilers (Arbor Acres, 1 D of age) were randomly allocated into 4 groups with 8 replicates of 8 birds. Dietary treatments were supplemented with taurine at the level of 0.00, 2.50, 5.00, and 7.50 g/kg of the diet (denoted as CON, TAU1, TAU2, TAU3, respectively). The BW gain from 1 to 21 D and from 22 to 42 D were all increased linearly (linear, P

The rapid renewal and repair of the intestinal mucosa are based on intestinal stem cells (ISC), which are located at the crypt bottom. Paneth cells are an essential component in the crypt, which served as the niche for ISC development. However, in the chicken, how the function of Paneth cells changes during intestinal inflammation is unclear and is the key to understand the mechanism of mucosal repair. In the present study, 36 HyLine White chickens (7 d of age, n = 6) were randomly divided into 1 control and 5 lipopolysaccharide (LPS) injection groups. The chickens were injected (i.p.) with PBS in the control group, however, were injected (i.p.) with LPS (10 mg/kg BW) in the LPS injection groups, which would be sampled at 5 time points (1 h postinjection [hpi], 2 hpi, 4 hpi, 6 hpi, and 8 hpi). Results showed that tumor necrosis factor-α mRNA transcription in duodenal tissue increased gradually since 1 hpi, peaked at 4 hpi, and then reduced remarkably, indicating that 4 hpi of LPS was the early stage of intestinal inflammation. Meanwhile, the MUC2 expression in duodenal tissue was dramatically reduced since 1 hpi of LPS. The ISC marker, Lgr5 and Bmi1, in the duodenal crypt were reduced from 1 hpi to 4 hpi and elevated later. Accordingly, the hydroethidine staining showed that the reactive oxygen species level, which drives the differentiation of ISC, in the duodenal crypt reduced obviously at 1 hpi and recovered gradually since 4 hpi. The analysis of Paneth cells showed that many swollen mitochondria appeared in Paneth cells at 4 hpi of LPS. Meanwhile, the Lysozyme transcription in the duodenal crypt was substantially decreased since 1 hpi of LPS. However, the Wnt3a and Dll1 in duodenal crypt decreased at 1 hpi of LPS, then increased gradually. In conclusion, Paneth cells were impaired at the early stage of intestinal inflammation, then recovered rapidly. Thus, the ISC activity was reduced at first and recovery soon.

Summary The larvae of the genus Baylisascaris can cause larva migrans in mammals and birds. This study investigated the larval migration of Baylisascaris potosis, the roundworm of kinkajou (Potos flavus), in chickens and the associated clinical manifestations of the host. Thirty-six 3-week-old chickens divided into 6 groups were orally inoculated with 3,000 B. potosis eggs/chick. Each group of chicken was necropsied at days 1, 2, 3, 7, 30 and 90 PI (post inoculation), and the number of larvae in various organs were counted until day 90 PI. No clinical signs were observed in chickens during the study. Larvae were detected from the liver, lungs or breast-muscles of 13/36 (36.1%) chickens. The mean total number of larvae in the liver, lungs and breast-muscles at days 1, 2, 3, 7, 30 and 90 PI were 0.34, 0.17, 1.66, 1.01, 0.17 and 0, respectively. No larvae were found in the brain, eyes, hid-limb muscles, heart, kidneys and spleen. Although infectivity of larvae in egg-inoculated chickens was low, the present study demonstrated that B. potosis larvae can migrate in chickens tissues up to day 30 PI. The result suggests that chickens can serve as a paratenic host for B. potosis and may underline a public health importance of B. potosis infection as a potential foodborne disease in humans.

The recent interest in plant-based foods has brought upon the need to develop novel structures using plant-based proteins. However, there is still room for improvement in the development of plant-based meat and cheese alternatives. The rheological properties of self-assembled zein networks were examined to evaluate potential in animal protein replacement. These plant-based protein networks were compared to gluten networks (a common ingredient in current plant-based products), chicken muscle tissue, and cheddar cheese. All samples were analyzed using temperature, amplitude, and frequency sweeps at different time points. Zein networks exhibited unique viscous behaviour (in line with that of an entangled polymer solution), in each amplitude, frequency and temperature sweeps, however only when freshly formed. The results suggest that the bonds and interactions responsible for strengthening zein networks need at least 24 h to fully form. Analysis of the secondary structure by FTIR revealed that zein undergoes a structural reorganization from intermolecular to intramolecular β-sheets during this time, but the substantial content of α-helix structures remains unchanged. Overall, different aspects of zein network rheological behaviour can be compared to either chicken breast, or cheddar cheese, presenting opportunities for zein in plant-based food structuring., Graphical abstract Image 1, Highlights - The rheological behavior of zein networks is investigated - Rheological properties of zein networks stabilized after 24 h of storage - Zein networks and cheddar cheese are brittle and demonstrate melting behavior - Zein networks and chicken muscle tissue demonstrate similar elastic properties

Objective: Follicle selection is an important process in chicken egg laying. Among several small yellow (SY) follicles, the one exhibiting the highest expression of follicle stimulation hormone receptor (FSHR) will be selected to become a hierarchal follicle. The role of lncRNA, miRNA and other non-coding RNA in chicken follicle selection is unclear. Methods: In this study, the whole transcriptome sequencing of SY follicles with different expression levels of FSHR in Jining Bairi hens was performed, and the expression of 30 randomly selected mRNAs, lncRNAs and miRNAs was validated by quantitative real-time polymerase chain reaction. Preliminary studies and bioinformatics analysis were performed on the selected mRNA, lncRNA, miRNA and their target genes. The effect of identified gene was examined in the granulosa cells of chicken follicles. Results: Integrated transcriptomic analysis on chicken SY follicles differing in FSHR expression revealed 467 differentially expressed mRNA genes, 134 differentially expressed lncRNA genes and 34 differentially expressed miRNA genes, and sosondowah ankyrin repeat domain family member A (SOWAHA) was the common target gene of three miRNAs and one lncRNA. SOWAHA was mainly expressed in small white (SW) and SY follicles and was affected by follicle stimulation hormone (FSH) treatment in the granulosa cells. Knockdown of SOWAHA inhibited the expression of Wnt family member 4 (Wnt4) and steroidogenic acute regulatory protein (StAR) in the granulosa cells of prehierarchal follicles, while stimulated Wnt4 in hierarchal follicles. Overexpression of SOWAHA increased the expression of Wnt4 in the granulosa cells of prehierarchal follicles, decreased that of StAR and cytochrome P450 family 11 subfamily A member 1 in the granulosa cells of hierarchal follicles and inhibited the proliferation of granulosa cells. Conclusion: Integrated analysis of chicken SY follicle transcriptomes identified SOWAHA as a network gene that is affected by FSH in granulosa cells of ovarian follicles. SOWAHA affected the expression of genes involved in chicken follicle selection and inhibited the proliferation of granulosa cells, suggesting an inhibitory role in chicken follicle selection.

Fatty liver hemorrhagic syndrome (FLHS), usually occurring in hens in the late laying period, can lower egg yield and even cause death. Similar to nonalcoholic fatty liver disease in humans, FLHS may begin with simple hepatic steatosis. It is known that gut microbiota is important to the development of nonalcoholic fatty liver disease, but the relationship between FLHS and gut bacteria remains unclear. In this study, bacteria compositions in the ileum and cecum were determined by 16S rDNA sequencing analysis in the groups of hens with vs. without hepatic steatosis (average liver fat contents were 0.34 mmol/g protein or 2.6% vs. 0.84 mmol/g protein or 6.0%). These hens were in the 2 tails of the liver fat content distribution with each tail accounting for 10% of the test population containing 90 66-wk-old healthy Rhode Island Red laying hens raised under routine feeding regimen. The results showed that liver weight but not the weights of the body, heart and abdominal fat were significantly different between the groups. Moreover, bacterial diversity was not significantly different between the groups, but bacterial diversity in the cecum was higher than that in the ileum. Proteobacteria was the most dominant phylum in the ileum, whereas Proteobacteria, Firmicutes, and Bacteroidetes were the most dominant phyla in the cecum. Some bacteria were common to the ileum and cecum, but some were unique. Furthermore, some bacteria were significantly different in abundance between the groups, that is the group without hepatic steatosis had more bacteria inhibiting host energy absorption or benefiting intestinal health, whereas the group with hepatic steatosis had more conditionally pathogenic or harmful bacteria. In conclusion, hepatic steatosis in laying hens is associated with intestinal bacterial composition (bacterial abundance) but not bacterial diversity. The identified common and unique bacteria are related to the functional characteristics of the ileum and cecum. The decrease of beneficial bacteria and the increase of harmful bacteria in the intestine of laying hens may increase FLHS incidence by promoting hepatic steatosis.

Background Our previous study found that chicken KLF7 was an important regulator in formation of adipose tissue. In the present study, we analyzed the association for DNA methylation in chicken KLF7 with its transcripts of abdominal adipose tissue and blood metabolic indicators. Results The KLF7 transcripts of the adipose tissue of Chinese yellow broilers were associated with age (F = 6.67, P = 0.0035). In addition, the KLF7 transcripts were negatively correlated with blood glucose levels (r = − 0.61841, P = 0.0140). The DNA methylation levels of 26 CpG loci in the chicken KLF7 promoter and Exon 2 were studied by Sequenom MassArray. A total of 22 valid datasets were obtained. None of them was significantly different in relation to age (P > 0.05). However, the DNA methylation levels in the promoter were lower than those in Exon 2 (T = 40.74, P KLF7 transcripts and blood high-density lipoprotein levels, respectively, and many CpG loci were correlated with each other (P KLF7 transcripts (P KLF7 transcripts was built, and the contribution of z1 to the variation on KLF7 transcripts was 34.29%. Conclusions In conclusion, the KLF7 transcripts of chicken abdominal adipose tissue might be inhibited by DNA methylation in the promoter, and it might be related to the DNA methylation level of PCpG6.

Background and Aim: Salmonella species often cause systemic health problems in poultry flocks, sometimes including nervous systems manifestations. This impact of Salmonella has rarely been studied. This study aimed to define an alternative pathogenic pathway for Salmonella spp. invasion of brain tissue in chicken flocks. Brain infection produces neurological manifestations; Salmonella strains isolated from brain tissue showed the presences of two virulence genes. Confirmation of the pathway of isolates from intestinal mucosa through the blood–brain barrier was attained using experimental infections in specific pathogen-free (SPF)-day-old chicks through two routes of inoculation. Materials and Methods: Isolation of Salmonella spp. from five chicken flocks that showed signs of the central nervous system (CNS) effects were isolated. Isolates were characterized by serotyping, and antimicrobial assays. In addition, virulence profiles were described using detection of virulence plasmid spvC, and Salmonella plasmid sopB. A pathogenicity study of isolates in specific pathogen-free (SPF)-day-old chicks through oral and intracerebral administration performed, and experimental infection in SPF embryonated chicken eggs through intra-yolk and intra-allantoic administration was investigated. Supporting histopathology and immunohistopathology against Salmonella antigen in brain tissue were performed for flock and experimental infections. Results: Three serotypes of Salmonella were isolated from the brains of five flocks (two Salmonella Virchow, two Salmonella Kentucky, and one Salmonella Enteritidis isolates). Phage related gene sopB and plasmid-mediated operon spvC were identified in all isolated strains. The Salmonella strains were re-isolated and identified from the brain and internal organs of post-experimental infected chicks. Infected chicks showed nervous manifestations associated with Salmonella infection. The presence of positively stained Salmonella antigen in brain tissues indicates penetration of the blood–brain barrier by the Salmonella species. Conclusion: Our results indicate that some virulent systemic strains of Salmonella spp. can induce CNS manifestations in chicken hosts.

Sheep Red Blood Cells (SRBC) are commonly employed by scientists to address humoral immune responses in poultry. While SRBC are closely related to the study of humoral immunity in poultry, the initial purpose of much research did not focus on the mechanisms involved. Here, we provide a qualitative approach, and utilize scientometric techniques, including trend analyses, scientific collaborations and mapping, and word co-occurrence evaluations, to summarize the role of SRBC in the poultry studies. First, a search strategy on Web of Science (WoS) was conducted to find publications that included SRBC in the poultry studies. Publications were partitioned into four categories: nutrition, genetics, microbiology, and physiology. For scientometric evaluation, scientific maps and networks were produced to clarify the occurrence of SRBC in the poultry studies. Data used were included 702 publications over a period of 50 years (1968-2018) that were retrieved from the WoS database. About 95% of the publications were published in English language. Indigenous, experimental and commercial chickens, quail, and medicinal plants field/topics were the main subjects of publications. In recent years, authors have used SRBC to study humoral immune response as a secondary aim of their research, especially when poultry production/performance was studied. This was especially the case in recent decades for studies in poultry nutrition. Analysis of keywords co-occurrence showed that the phrase SRBC mostly occurred with chickens, immune response, and especially with broilers. Moreover, the “medicinal plants” is becoming important especially for research on broilers, and the reduced use of antibiotics in feed. Consequently, in addition to studying the medicinal plants, finding antibiotic replacements, and/or growth performance in the birds, humoral immunity is suggested to be investigated using SRBC. Moreover, inter-disciplinary studies with the cooperation of scientists from agriculture, veterinary, immunology, biochemistry and molecular biology, and toxicology will develop in the future.

The plastic component bisphenol A (BPA) impairs reproductive organ development in various experimental animal species. In birds, effects are similar to those caused by other xenoestrogens. Because of its endocrine disrupting activity, BPA is being substituted with other bisphenols in many applications. Using the chicken embryo model, we explored whether the BPA alternatives bisphenol AF (BPAF), bisphenol F (BPF), and bisphenol S (BPS) can induce effects on reproductive organ development similar to those induced by BPA. Embryos were exposed in ovo from embryonic day 4 (E4) to vehicle, BPAF at 2.1, 21, 210, and 520 nmol/g egg, or to BPA, BPF, or BPS at 210 nmol/g egg and were dissected on embryonic day 19. Similar to BPA, BPAF and BPF induced testis feminization, manifested as eg testis-size asymmetry and ovarian-like cortex in the left testis. In the BPS-group, too few males were alive on day 19 to evaluate any effects on testis development. We found no effects by any treatment on ovaries or Müllerian ducts. BPAF and BPS increased the gallbladder-somatic index and BPAF, BPF and BPS caused increased embryo mortality. The overall lowest-observed-adverse-effect level for BPAF was 210 nmol/g egg based on increased mortality, increased gallbladder-somatic index, and various signs of testis feminization. This study demonstrates that the BPA replacements BPAF, BPF, and BPS are embryotoxic and suggests that BPAF is at least as potent as BPA in inducing estrogen-like effects in chicken embryos. Our results support the notion that these bisphenols are not safe alternatives to BPA.

In this study, a strain of Trichosporon was isolated from white pseudomembranes and ulcers formed on mucous membranes of pigeon bursas and was identified through gene sequencing. Bacteriostatic actions of Acorus gramineus, Sophora flavescens, Polygonum hydropiper, and Chinese herbal mixture on this species were explored in vitro, and the minimum inhibitory concentration of herbal medicines against Trichosporon was determined through microdilution method. Therapeutic effects of herbal medicines on chickens infected by Trichosporon were studied, whose results showed that minimum inhibitory concentration of A. gramineus was 32 μg/μL, that of S. flavescens was 2 μg/μL, that of P. hydropiper was 120 μg/μL, and that of Chinese herbal mixture was 36 μg/μL. Antibacterial effects of S. flavescens were the best. In accordance with animal experiments, therapeutic effects of Chinese herbal medicines on infected chickens were better than those of fluconazole. The mortality rate of the Chinese herbal medicine treatment group was 33.33%, that of the fluconazole treatment group was 46.67%, and that of the Chinese medicine protection group was 23.33%. The longer the time of Chinese medicine treatments was, the better the treatment effects would be. Glutamic oxaloacetylase values of the serum and liver in the Chinese herbal medicine treatment group were both significantly lower than those of the nontreatment group. From the results, it can be seen that A. gramineus, S. flavescens, P. hydropiper, and Chinese herbal mixture have certain inhibitory effects on Trichosporon spp. Chinese herbal medicine protections in advance could reduce Trichosporon infections.

Avian leucosis (AL) is a disease characterized by tumors and is caused by the avian leukosis virus (ALV). Because of the high variability of viruses and complex pathogenic mechanisms, screening and breeding J subgroup of ALV (ALV-J) resistant avian breeds is one of the strategies for prevention and treatment of AL, thus screening of significant immune markers is needed to promote the development of disease-resistant breeds. In this study, data-independent acquisition (DIA) technology was used to detect the DEPs of three breeds of chicken according to different comparison to investigate the potential markers. Results showed special DEPs for spleen development of each breed were detected, such as PCNT, DDB2, and ZNF62. These DEPs were involved in intestinal immune network used in production of IgA signaling pathways and related to immune response which can be used as potential markers for spleen development in different breeds. The DEPs such as RAB44 and TPN involved in viral myocarditis, transcriptional misregulation in cancer, and tuberculosis can be used as potential markers of spleen immune response after ALV-J infection in chickens. Pair-wise analysis was performed for the three breeds after the infection of ALV-J. The proteins such as RFX1, TAF10, and VH1 were differently expressed between three breeds. These DEPs involved in antigen processing and expression, acute myelogenous leukemia, and viral carcinogenesis can be used as potential immune markers after ALV-J infection of different genetic backgrounds. The screening of potential markers at protein level provides a strong theoretical research basis for disease resistance breeding in poultry.

Across the world, many commercial poultry flocks and captive birds are threatened by infection with Aspergillus fumigatus. Susceptibility to aspergillosis varies among birds; among galliform birds specifically, morbidity and mortality rates seem to be greater in turkeys than in chickens. Little is known regarding the features of avian immune responses after inhalation of Aspergillus conidia, and to date, scarce information on inflammatory responses during aspergillosis exists. Thus, in the present study, we aimed to improve our understanding of the interactions between A. fumigatus and economically relevant galliform birds in terms of local innate immune responses. Intra-tracheal aerosolization of A. fumigatus conidia in turkey and chicken poults led to more severe clinical signs and lung lesions in turkeys, but leukocyte recovery from lung lavages was higher in chickens at 1dpi only. Interestingly, only chicken CD8+ T lymphocyte proportions increased after infection. Furthermore, the lungs of infected chickens showed an early upregulation of pro-inflammatory cytokines, including IL-1β, IFN-γ and IL-6, whereas in turkeys, most of these cytokines showed a downregulation or a delayed upregulation. These results confirmed the importance of an early pro-inflammatory response to ensure the development of an appropriate anti-fungal immunity to avoid Aspergillus dissemination in the respiratory tract. In conclusion, we show for the first time that differences in local innate immune responses between chickens and turkeys during aspergillosis may determine the outcome of the disease. Lay summary Aspergillus fumigatus infection may cause mortality in poultry, depending on species sensitivity. This study confirms the earlier activation of chickens' pro-inflammatory effectors to control Aspergillus dissemination, whereas turkeys' immune response enables the exacerbation of lung lesions.

Background The 3D8 single chain variable fragment (scFv) is a mini-antibody sequence that exhibits independent nuclease activity against all types of nucleic acids. In this research, crossing a 3D8 scFv G1 transgenic rooster with wild-type hens produced 3D8 scFv G2 transgenic chickens to evaluate suppression of viral transmission. Result The transgenic chickens were identified using genomic PCR and immunohistochemistry. To evaluate Newcastle disease virus (NDV) protection conferred by 3D8 scFv expression, transgenic, non-transgenic, and specific pathogen-free (SPF) chickens were challenged with virulent NDV by direct injection or aerosol exposure. The three groups of chickens showed no significant differences (p Conclusions Our finding suggest that the 3D8 scFv could be a useful tool to prevent chickens from spreading NDV and control virus transmission.

Public discourse around “greenhouse gases” (GHG) has led to the application of life-cycle assessments to ascertain the “global warming potential” of human activities. Life-cycle assessments applied to agricultural systems typically do not consider positive contributions (i.e., fixation of atmospheric carbon dioxide [CO2]) or consider complex interrelationships among commodities within the larger agricultural sector. The purpose of this article is to present an argument for a paradigm shift and that poultry production should be considered as a value-adding activity within modern crop production systems for GHG foot-printing purposes. To this end, a case study based on 2018 production data is presented where poultry production (chicken and eggs) was contextualized as a sub-component of wheat and corn production in the Canadian provinces of Alberta and Ontario, respectively. Total GHG footprint was calculated to be 3.05 and 3.29 million tonnes (MT) of CO2 equivalent (eq) for Alberta wheat and Ontario corn production, respectively. The GHG footprint of chicken production was calculated to be 0.39 and 1.38 MT CO2 eq in Alberta and Ontario, respectively. The GHG footprint of egg production calculated to be 0.12 and 0.47 MT of CO2 eq in Alberta and Ontario, respectively. When carbon (C) fixation as crop biomass is included in the scenario, the combined crop–poultry system C balance in 2018 favored net fixation of 40.70 and 35.15 MT of CO2 eq in Alberta and Ontario, respectively. The calculated total GHG footprint of poultry production in Alberta and Ontario corresponded to only 1.2 and 5.5% of the calculated total net CO2 fixation of their respective cropping systems. This case study demonstrates that by failing to acknowledge real world estimates of C fixation by crop biomass, GHG foot-printing exercises largely misrepresent reality and can thus perpetuate faulty assumptions about the environmental footprint of animal agriculture. The authors propose that the calculations presented herein provide grounds to postulate the hypothesis that modern, integrated crop-livestock agricultural systems in Canada (and elsewhere) act as net sinks for atmospheric CO2.

Background and Aim: Testis (T) and epididymis (E) are waste from the abattoir that is rarely used. In fact, both organs contain important chemicals needed for spermatogenesis (e.g., hormones, proteins, and other molecules). Therefore, administration of a combination of testis and epididymis (CTE) extracts may activate androgen receptors (AR) and protein kinase A (PKA) molecules that play a prominent role in spermatogenesis. We, therefore, aimed at investigating the influence of the CTE extracts on the concentration of AR and PKA in male chicken. Materials and Methods: This study used a completely randomized design with four treatment groups (K0, K1, K2, and K3) and five replications per group. K0 is a control group that received 1 mL normal saline, whereas K1, K2, and K3 are the test groups that received 1, 2, and 3 mL of CET extracts, respectively. Twenty male chickens (strain: broiler Mb 89), 3 weeks of age, weighing 500-700 g were used. We administered the injections in a 13-day period and on the 14th day; we collected and processed blood samples as serum to measure the AR and PKA concentrations using commercial chicken AR and PKA enzyme-linked immunosorbent assay kits, respectively. We performed analyses by analysis of variance using SPSS 20.0. Results: The AR concentrations in K1, K2, and K3 groups increased by 4.26%, 10.97%, and 28.04%, respectively, compared to the K0 (control group). However, this increase was not significantly different between the groups (p>0.05). Moreover, the PKA concentrations increased by 2.97%, 2.60%, and 4.08% in K1, K2, and K3 groups, respectively, compared to the control group. However, this increase was not significantly different between the groups as well (p>0.05). Conclusion: The CTE extracts tended to increase the AR and PKA concentrations even though it is not significant. Therefore, it needs further study when using the CTE extracts for spermatogenesis in male chicken.

Antimicrobial resistance (AMR) is a major public health threat worldwide. The main objective of this study was to compare AMR in Campylobacter from broiler chickens raised on Canadian farms and their products in different geographical regions of Canada. To do this, antimicrobial susceptibility results from isolates of Campylobacter recovered from a national microbiological baseline study conducted in federally registered establishments and in the retail marketplace were analyzed. Among 1460 isolates tested, 774 (53%) were resistant to at least one antimicrobial, with a predominance of three profiles: tetracycline (39%), quinolone–tetracycline (6.6%), and quinolones only (3.5%). The results showed no significant difference in the frequency of resistant profiles (p ≥ 0.05) among the isolates originating from different points in the food processing chain at slaughterhouses and in retail establishments. This suggests that AMR observed in Campylobacter isolates from raw chicken at retail originated further upstream in the system. A difference in the frequency of certain resistance profiles was observed between the regions of Canada. For instance, in British Columbia, there was more resistance to quinolones, while in Ontario and Quebec, Campylobacter isolates were more resistant to tetracyclines, macrolides, ketolides, and lincosamides. Comparison of AMR data from this study with those from the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS) did not show any significant difference and provides evidence that CIPARS produces nationally representative resistance results.

This study aimed to investigate the effects of selenide chitosan sulfate (Se-CTS-S) on glutathione (GSH) system in hepatocytes and chickens. Chitosan, sodium selenite (Na2SeO3), selenide chitosan, chitosan sulfate (CTS-S), and Se-CTS-S were added to the culture medium and the basal diets; glutathione peroxidase (GSH-Px) activity, GSH content, total antioxidant capacity (T-AOC), and mRNA levels of cellular GPx (GPx-1) and phospholipid hydroperoxide GPx (GPx-4) in vivo and in vitro were determined. The results showed that Se-CTS-S increased (P < 0.05) GPx-1 and GPx-4 mRNA levels in hepatocytes and livers, and GSH-Px activity, GSH content, and T-AOC in the medium, hepatocytes, plasma, and livers compared with the control and chitosan treatments. Compared with CTS-S, Se-CTS-S treatments increased (P < 0.05) GPx-1 and GPx-4 mRNA levels in hepatocytes and livers, and GSH-Px activity, GSH content, and T-AOC capacity in the medium, hepatocytes, and livers. Compared with Na2SeO3 and CTS-Se, Se-CTS-S increased (P < 0.05) GPx-1 mRNA levels in hepatocytes and livers, GPx-4 mRNA levels in hepatocytes and livers, GSH-Px activity in the medium, hepatocytes, and livers, GSH contents in plasma and livers, and T-AOC in the medium, plasma, and livers. Thus, Se-CTS-S showed better biological activity that mainly benefited from the synergistic effects of Se and sulfate on GSH system.

International audience; AbstractContinued reports of infections with infectious bronchitis virus (IBV) variants have occurred since its first isolation in the 1930s. Currently, QX-like IBVs are the predominant circulating genotype around the world. Here, the pathogenicity of QX-like IBV strain SD was characterized in chickens at different ages of exposure to the virus, and the protection efficacy of available vaccine combinations against IBV was evaluated. The results revealed that QX-like IBV strain SD was severely pathogenic in chickens, causing respiratory, urinary and reproductive infections, irrespective of age, based on clinical observations, viral distribution in tissues and a ciliostasis study. Severe respiratory signs, tracheal cilia injury, nephritis and abnormal development of the oviduct and ovarian follicles were evident throughout the experiment. A challenge experiment demonstrated that the homologous QX vaccine showed superior protection efficacy compared with other available vaccines, confirming the importance of IBV vaccine seed homology against the circulating IBV strains. Our findings aid an understanding of the pathogenicity of QX-like IBVs that may help to further control the infection.

Background Marek’s disease (MD) is a highly neoplastic disease primarily affecting chickens, and remains as a chronic infectious disease that threatens the poultry industry. Copy number variation (CNV) has been examined in many species and is recognized as a major source of genetic variation that directly contributes to phenotypic variation such as resistance to infectious diseases. Two highly inbred chicken lines, 63 (MD-resistant) and 72 (MD-susceptible), as well as their F1 generation and six recombinant congenic strains (RCSs) with varied susceptibility to MD, are considered as ideal models to identify the complex mechanisms of genetic and molecular resistance to MD. Results In the present study, to unravel the potential genetic mechanisms underlying resistance to MD, we performed a genome-wide CNV detection using next generation sequencing on the inbred chicken lines with the assistance of CNVnator. As a result, a total of 1649 CNV regions (CNVRs) were successfully identified after merging all the nine datasets, of which 90 CNVRs were overlapped across all the chicken lines. Within these shared regions, 1360 harbored genes were identified. In addition, 55 and 44 CNVRs with 62 and 57 harbored genes were specifically identified in line 63 and 72, respectively. Bioinformatics analysis showed that the nearby genes were significantly enriched in 36 GO terms and 6 KEGG pathways including JAK/STAT signaling pathway. Ten CNVRs (nine deletions and one duplication) involved in 10 disease-related genes were selected for validation by using quantitative real-time PCR (qPCR), all of which were successfully confirmed. Finally, qPCR was also used to validate two deletion events in line 72 that were definitely normal in line 63. One high-confidence gene, IRF2 was identified as the most promising candidate gene underlying resistance and susceptibility to MD in view of its function and overlaps with data from previous study. Conclusions Our findings provide valuable insights for understanding the genetic mechanism of resistance to MD and the identified gene and pathway could be considered as the subject of further functional characterization.

Background Avian eggs have a proteinaceous cuticle. The quantity of cuticle varies and the deposition of a good cuticle in the uterus (Shell-gland) prevents transmission of bacteria to the egg contents. Results To understand cuticle deposition, uterus transcriptomes were compared between hens with i) naturally good and poor cuticle and, ii) where manipulation of the hypothalamo-pituitary-gonadal-oviduct axis produced eggs with or without cuticle. The highest expressed genes encoded eggshell matrix and cuticle proteins, e.g. MEPE (OC-116), BPIFB3 (OVX-36), RARRES1 (OVX-32), WAP (OVX-25), and genes for mitochondrial oxidative phosphorylation, active transport and energy metabolism. Expression of a number of these genes differed between hens laying eggs with or without cuticle. There was also a high expression of clock genes. PER2, CRY2, CRY1, CLOCK and BMAL1 were differentially expressed when cuticle deposition was prevented, and they also changed throughout the egg formation cycle. This suggests an endogenous clock in the uterus may be a component of cuticle deposition control. Cuticle proteins are glycosylated and glycosaminoglycan binding genes had a lower expression when cuticle proteins were deposited on the egg. The immediate early genes, JUN and FOS, were expressed less when the cuticle had not been deposited and changed over the egg formation cycle, suggesting they are important in oviposition and cuticle deposition. The uterus transcriptome of hens with good and poor cuticle deposition did not differ. Conclusions We have gained insights into the factors that can affect the production of the cuticle especially clock genes and immediate early genes. We have demonstrated that these genes change their expression over the period of eggshell formation supporting their importance. The lack of differences in expression between the uterus of hens laying eggs with the best and worse cuticle suggest the genetic basis of the trait may lie outside the oviduct.

In the present study we hypothesize that aquaporin 4 (AQP4) expression in the chicken oviduct would change during a pause in egg laying that was induced by fasting. Accordingly, the aim of this investigation was to examine the AQP4 mRNA and protein expression, and immunolocalization in the chicken oviduct during the course of regression. The experiment was carried out on laying hens subjected to a pause in laying that was induced by food deprivation for 5 days. Control hens were fed ad libitum. The birds were sacrificed on day 6 of the experiment and all segments of the oviduct were isolated, including the infundibulum, magnum, isthmus, shell gland, and vagina. Subsequently, the gene and protein expressions of AQP4 in the tissues were tested by real-time PCR and Western blot, respectively. The relative mRNA expression of AQP4 was the highest in the infundibulum and vagina and the lowest, and least detectable, in the magnum. The level of AQP4 protein was the highest in the infundibulum and the lowest in the magnum. Fasting resulted in a decrease of the AQP4 mRNA expression ( P 0 . 001 ) in the infundibulum, a decrease in protein abundance ( P 0 . 01 ) in the shell gland, and an increase in protein level ( P 0 . 001 ) in the vagina. Immunohistochemistry demonstrated tissue- and cell-dependent localization of AQP4 protein in the oviductal wall. The intensity of staining was as follows: the infundibulum > shell gland > vagina ≥ isthmus ≫ magnum. In the control hens, the immunoreactivity for AQP4 in the vagina was similar, whereas in other oviductal segments, the immunoreactivity was stronger when compared with the chickens subjected to a pause in laying. In summary, these findings suggest that the AQP4 is an essential protein involved in the regulation of water transport required to create a proper microenvironment for fertilization and egg formation in the hen oviduct.

Highly sensitive detection of pathogens is effective for screening meat during quarantine inspection and export. The “micro-amount of virion enrichment technique” (MiVET) was recently developed, which is a new method combining virus concentration with immunomagnetic beads and simple RNA extraction with sodium dodecyl benzenesulfonate (SDBS) for the specific and sensitive detection of avian influenza viruses (AIVs). AIV subtypes H3N2 and H4N2 were used to spike the surface of chicken breast meat samples. The modified MiVET protocol was tested by comparing it against three different homogenate preparation conditions, as well as in samples with added α-amylase and collagenase to digest inhibitors. The performance of the modified MiVET was evaluated by real-time RT-PCR assay targeting the matrix gene. Compared with conventional RNA extraction, the modified MiVET reproducibly concentrated AIVs in chicken meat samples with 100–1000-fold improvement by 60 s-hand homogenization. The 30 s- and 60 s-stomacher homogenizations resulted 100-fold and 10–100-fold improvement, respectively. The modified MiVET required

This study was performed to investigate the effects of lignocellulose supplementation (LS) on performance parameters, egg quality, aerobic bacterial load of eggshell, serum biochemical parameters, and jejunal histomorphological traits of laying hens between 18 and 38 wk of age. A total of 640 pullets at 16 wk of age were allotted to 4 treatment groups as 0 kg (control, CONT), 0.5 kg, 1 kg, and 2 kg LS per ton of feed. Body weight (BW), daily feed intake, egg production (EP), egg weight (EW), and efficiency of feed utilization (EF) were determined as the mean of each 3-wk period between 18 and 38 wk of age. Laying hens in the 1 kg LS group had a higher BW mean (1632.1 g, P < 0.001). the highest mean value of EP and EW were observed in 1 kg LS group (81.8% and 57.3 g, respectively), whereas the lowest values were found in the 2 kg LS group (78.6% and 54.4 g, respectively, P < 0.001). the mean of EF was the lowest in the 1 kg LS group (2.72, P < 0.001). There was a decline in eggshell breaking strength and eggshell thickness in the 2 kg LS, when compared with the 0.5 and 1 kg LS groups (P < 0.001). the total aerobic bacterial load of the eggshell was the lowest in the 1 kg LS group (4.7 log10 cfu/mL). the level of aspartate amino transferase and alanine amino transferase showed an incre-ment in both the CONT and 2 kg LS groups (P < 0.001). the high level of LS (2 kg per ton of feed) caused a decline in the levels of IgY, IgA and IgM, when compared to the 0.5 and 1 kg LS groups (P < 0.001). Laying hens in 0.5 and 1 kg LS groups had longer villus height (1335.9 mm) in the jejunum than the others (P < 0.001). These find-ings showed that the 1 kg LS per ton of feed improved EP and EW, eggshell quality, immunoglobulin levels and intestinal morphology, and decreased the total aerobic bacterial load., Global Nutritech Biotechnology LLC (Richmond, VA, USA), The authors would like to thank Global Nutritech Biotechnology LLC (Richmond, VA, USA) for their financial support.

Large ovarian follicles are primary characteristics of oviparous species. The development of such follicles is crucially governed by strict intrinsic complex regulation. Many aspects of the genetic basis of this regulation remain obscure. To identify the dominant genes controlling follicular development in the chicken, growing follicles (400–1,600 μm in diameter) were selected for RNA sequencing and bioinformatics analysis. Comparing the 400-μm follicles with 800-μm follicles identified a total of 3,627 differentially expressed genes (1,792 upregulated and 1,835 downregulated genes). Comparing the 400-μm follicles with 1,600-μm follicles revealed 9,650 differentially expressed genes (including 4,848 upregulated and 4,802 downregulated genes). Comparing 800-μm with 1,600-μm follicles revealed a total of 6,779 differentially expressed genes (3,427 upregulated and 3,352 downregulated genes). Transcriptome analysis revealed that genes related to the extracellular matrix–receptor interactions, steroid biosynthesis, cell adhesion, and phagosomes displayed remarkable differential expressions. Relative to 400-μm follicles, collagen content, production of steroid hormones, cell adhesion, and phagocytic factors were significantly increased in the 1,600-μm follicles. This study identifies the dominant genes involved in the promotion of follicular development in oviparous vertebrates and represents the extraordinary gene regulation pattern related to development of the growing follicles in poultry.

Background Globally, chicken is known to be a reservoir for the spread of antimicrobial resistance genes to humans. In Nigeria, antimicrobial drugs are readily accessible for use in poultry production, either for preventive or therapeutic purposes. Extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-EC) are transmissible to humans because of their zoonotic potentials. People working very closely with chickens either on farms or markets are at greater risk. The aim of this study was to investigate the prevalence and zoonotic transmission of ESBL-EC among poultry-workers, chickens, and poultry environments in Abuja, Nigeria. Methods We conducted a cross-sectional study among workers, chickens and poultry environment in selected farms/chicken markets in Abuja. Stool, faecal, and environmental samples were collected from apparently healthy workers, chickens, and farm/market environments from December 2018 to April 2019. Data were collected electronically using an open data kit (ODK) installed on a Smartphone. Antimicrobial resistance was determined using broth micro-dilution methods against a panel of 14 antimicrobial agents. We carried out the phenotypic and genotypic characterization of the isolates. Data were analyzed by computing frequencies, proportions and spearman’s correlation (ρ). Results Of 429 samples, 26.8% (n = 115) were positive for Escherichia coli (E. coli). Of the 115 E. coli isolates, 32.2% (n = 37) were confirmed ESBL producers by phenotypic characterization. Prevalence of ESBL-EC was highest among both poultry-workers (37.8%; n = 14) and chickens (37.8%; n = 14) followed by the environment (24.3%; n = 9). Both human and chicken isolates showed similar patterns of multidrug resistance to tested antimicrobials with a positive correlation (ρ = 0.91). Among ESBL producers, we observed the dissemination of blaCTX-M (10.8%; n = 4) genes. The coexistence of blaCTX-M-15 and blaTEM-1 genes was observed in 8.1% (n = 3) of the isolates, out of which (66.7%; n = 2) were chicken isolates from the farm, while a single human isolate was from the chicken market. Conclusions ESBL-EC isolates were prevalent amongst apparently healthy individuals, chickens and the poultry farm/market environment in Abuja. It is important to educate healthcare workers that people in proximity with poultry are a high-risk group for faecal carriage of ESBL-EC, hence pose a higher risk to the general population for the spread of antimicrobial resistance.