Morphomolecular identification of heavy parasitic typhlitis in layer flocks: tissue response and cell-mediated reaction.

BACKGROUND: Heterakis gallinarum (H. gallinarum) is a common poultry parasite that can be found in the ceca of many gallinaceous bird species, causing minor pathology and reduced weight gain. Most infections go unnoticed in commercial flocks due to the dependence on fecal egg counts, which are prone to false-negative diagnoses. Furthermore, there is a lack of research on gastrointestinal nematodes that use molecular identification methods, which could be essential for rapid diagnosis and developing efficient control approaches. As a result, the study aimed to look at the cause of mortality in layer chickens induced by H. gallinarum in Egyptian poultry farms using morphological, ultrastructural, and molecular characterization. Histopathological, immunohistochemical, and cell-mediated immune responses from damaged cecal tissues were also examined. RESULTS: Seventy bird samples from ten-layer flocks of different breeds (Native, white, and brown layers) suffering from diarrhea, decreased egg output, and emaciation were collected. Cecal samples were collected from affected and non-affected birds and were examined for parasitic diseases using light and a scanning electron microscope. The mitochondrial cytochrome oxidase 1 (COX1) gene was used to characterize H. gallinarum. Our results showed that the collected nematodal worms were identified as H. gallinarum (male and female), further confirmed by COX1 gene amplification and sequence alignment. Gene expression analysis of the inflammatory markers in infected tissues showed a significant up-regulation of IL-2, IFN-γ, TLR-4, and IL-1ß and a significant down-regulation of the anti-inflammatory IL-10. The mRNA level of the apoptotic cas-3 revealed apoptotic activity among the H. gallinarum samples compared to the control group. CONCLUSIONS: Our results implemented the use of molecular methods for the diagnosis of Heterakis, and this is the first report showing the tissue immune response following infection in layers: upregulation of IL-1ß, IFN-γ, Il-2, and TLR-4, while down-regulation of anti-inflammatory IL-10 in cecal tissue, Cas-3 apoptotic activity and Nuclear factor-κB (NF-κB)activity with immunophenotyping of T-cells in Heterakis infected tissue.

Genetic Variation in Natural and Induced Antibody Responses in Layer Chickens.

Selection of livestock for disease resistance is challenging due to the difficulty in obtaining reliable phenotypes. Antibodies are immunological molecules that provide direct and indirect defenses against infection and link the activities of both the innate and adaptive compartments of the immune system. As a result, antibodies have been used as a trait in selection for immune defense. The goal of this study was to identify genomic regions associated with natural and induced antibodies in chickens using low-pass sequencing. Enzyme-linked immunosorbent assays were used to quantify innate (natural) antibodies binding KLH, OVA, and PHA and induced (adaptive) antibodies binding IBD, IBV, NDV, and REO. We collected plasma from four White Leghorn (WL), two White Plymouth Rock (WPR), and two Rhode Island Red (RIR) lines. Samples numbers ranged between 198 and 785 per breed. GWAS was performed within breed on data pre-adjusted for Line-Hatch-Sex effects using GCTA. A threshold of p = 10-6 was used to select genes for downstream annotation and enrichment analysis with SNPEff and Panther. Significant enrichment was found for the defense/immunity protein, immunoglobulin receptor superfamily, and the antimicrobial response protein in RIR; and the immunoglobulin receptor superfamily, defense/immunity protein, and protein modifying enzyme in WL. However, none were present in WPR, but some of the selected SNP were annotated in immune pathways. This study provides new insights regarding the genetics of the antibody response in layer chickens.

Phenotypic divergence between broiler and layer chicken lines is regulated at the molecular level during development.

BACKGROUND: Understanding the molecular underpinnings of phenotypic variations is critical for enhancing poultry breeding programs. The Brazilian broiler (TT) and laying hen (CC) lines exhibit striking differences in body weight, growth potential, and muscle mass. Our work aimed to compare the global transcriptome of wing and pectoral tissues during the early development (days 2.5 to 3.5) of these chicken lines, unveiling disparities in gene expression and regulation. RESULTS: Different and bona-fide transcriptomic profiles were identified for the compared lines. A similar number of up- and downregulated differentially expressed genes (DEGs) were identified, considering the broiler line as a reference. Upregulated DEGs displayed an enrichment of protease-encoding genes, whereas downregulated DEGs exhibited a prevalence of receptors and ligands. Gene Ontology analysis revealed that upregulated DEGs were mainly associated with hormone response, mitotic cell cycle, and different metabolic and biosynthetic processes. In contrast, downregulated DEGs were primarily linked to communication, signal transduction, cell differentiation, and nervous system development. Regulatory networks were constructed for the mitotic cell cycle and cell differentiation biological processes, as their contrasting roles may impact the development of distinct postnatal traits. Within the mitotic cell cycle network, key upregulated DEGs included CCND1 and HSP90, with central regulators being NF-κB subunits (RELA and REL) and NFATC2. The cell differentiation network comprises numerous DEGs encoding transcription factors (e.g., HOX genes), receptors, ligands, and histones, while the main regulatory hubs are CREB, AR and epigenetic modifiers. Clustering analyses highlighted PIK3CD as a central player within the differentiation network. CONCLUSIONS: Our study revealed distinct developmental transcriptomes between Brazilian broiler and layer lines. The gene expression profile of broiler embryos seems to favour increased cell proliferation and delayed differentiation, which may contribute to the subsequent enlargement of pectoral tissues during foetal and postnatal development. Our findings pave the way for future functional studies and improvement of targeted traits of economic interest in poultry.

Raillietina cesticillus infection causes reduced egg production in chickens in a windowless poultry house.

In a windowless poultry house raising layer chickens in Kanagawa prefecture, Japan, a slight increase in the mortality of chickens and a decrease in egg production were observed. Necropsy revealed numerous tapeworms and proglottids in chicken intestines. Histopathologically, gut-associated lymphoid tissues were observed in the lamina propria of the jejunum; however, no significant changes were observed in the other organs. Numerous hide beetles, Dermestes maculatus DeGeer, intermediate hosts of Raillietina cesticillus, were observed in the poultry house. Following a decline in beetle numbers, egg production increased and chicken mortality decreased. The life cycle of a tapeworm was easily established in a closed space, such as a windowless house, which led to severe infections.

Morphological and molecular characteristics of Plasmodium juxtanucleare in layer chicken from three districts of Yogyakarta, Indonesia.

Background and Aim: Blood parasite infections in poultry, such as Plasmodium, are a serious threat to the poultry industry due to their potential to cause economic losses. To date, there has been inadequate research on the morphological and molecular detection of the different Plasmodium species that infect poultry in Indonesia. Therefore, this study aimed to analyze the morphological and molecular characteristics of Plasmodium spp. and the several predisposing factors for Plasmodium infection in layer chickens from three districts of Yogyakarta, Indonesia. Materials and Methods: One hundred and five blood samples from layer chickens were collected from 13 farms located in three districts of Yogyakarta (Sleman, Bantul, and Kulon Progo) between September and November 2022. Blood samples were subjected to microscopic and polymerase chain reaction (PCR) analyses. Sequencing was performed using basic local alignment search tools to identify the nucleotide structure of cytochrome b. Phylogenetic analysis of Plasmodium was performed using the MEGA-X software. Results: Microscopic examination revealed that 17/105 positives (16.19%) were positive for blood parasite infection. Trophozoites, erythrocytic meronts, and microgametocytes of Plasmodium were found in blood samples. Based on the morphological examination, the species found in the samples was close to Plasmodium juxtanucleare. Polymerase chain reaction examination revealed that 21/60 samples were positive for Plasmodium (35%). The Plasmodium species identified from the sequenced samples were proven to be P. juxtanucleare. The P. juxtanucleare from Thailand was closely related to samples (99.64%-100%) with a genetic distance of 0%-1%. In addition, age, population, and cage type were not significantly associated with Plasmodium infection. Conclusion: Based on microscopic and PCR examinations, the Plasmodium species found in the three districts of Yogyakarta was P. juxtanucleare. The genetic distance between samples from the three districts of Yogyakarta was closely related (0%-1%) to P. juxtanucleare from Thailand and Japan. There was no correlation between Plasmodium infection and age, cage type, or population.

The abundance and diversity of antibiotic resistance genes in layer chicken ceca is associated with farm enviroment.

Industrialized layer chicken feedlots harbor complex environmental microbial communities that affect the enrichment and exchange of gut bacteria and antibiotic resistance genes (ARGs). However, the contribution of different environmental sources to the gut ARGs of layer chickens is not clear. Here, layer chicken gut and environmental samples (air, water, feed, cage, feather, maternal hen feces, uropygial glands) were collected during the early 3 month period before the laying of eggs, and the source and characteristics of the gut microorganisms and ARGs were analyzed by performing 16S rRNA and metagenomic sequencing. The results showed that the abundances of Bacteroidetes and Actinobacteria in cecum of layer chickens gradually increased, while that of Proteobacteria decreased with age, and the number and relative abundance of ARGs decreased significantly with age. On day 5, 57% of the layer chicken cecal ARGs were from feather samples, and 30% were from cage samples. Subsequently, the contribution of cage ARGs became progressively more prominent over time. At days 30 and 57, the contribution of cage ARGs to the chick cecal ARGs reached 63.3 and 69.5%, respectively. The bacterial community composition (especially the abundances of Klebsiella pneumoniae and Escherichia coli) was the major factor impacting the ARG profile. K. pneumoniae and E. coli were mainly transmitted from feathers to the layer chicken cecum, and the contribution rates were 32 and 3.4%, respectively. In addition, we observed the transmission of ARG-carrying bacteria (Bacteroides fragilis) from the cage to the gut, with a contribution rate of 11.5%. It is noteworthy that B. fragilis is an opportunistic pathogen that may cause diarrhea in laying hens. These results can provide reference data for the healthy breeding of layer chickens and the prevention and control of ARG pollution.

Restriction site-associated DNA sequencing technologies as an alternative to low-density SNP chips for genomic selection: a simulation study in layer chickens.

BACKGROUND: To reduce the cost of genomic selection, a low-density (LD) single nucleotide polymorphism (SNP) chip can be used in combination with imputation for genotyping selection candidates instead of using a high-density (HD) SNP chip. Next-generation sequencing (NGS) techniques have been increasingly used in livestock species but remain expensive for routine use for genomic selection. An alternative and cost-efficient solution is to use restriction site-associated DNA sequencing (RADseq) techniques to sequence only a fraction of the genome using restriction enzymes. From this perspective, use of RADseq techniques followed by an imputation step on HD chip as alternatives to LD chips for genomic selection was studied in a pure layer line. RESULTS: Genome reduction and sequencing fragments were identified on reference genome using four restriction enzymes (EcoRI, TaqI, AvaII and PstI) and a double-digest RADseq (ddRADseq) method (TaqI-PstI). The SNPs contained in these fragments were detected from the 20X sequence data of the individuals in our population. Imputation accuracy on HD chip with these genotypes was assessed as the mean correlation between true and imputed genotypes. Several production traits were evaluated using single-step GBLUP methodology. The impact of imputation errors on the ranking of the selection candidates was assessed by comparing a genomic evaluation based on ancestry using true HD or imputed HD genotyping. The relative accuracy of genomic estimated breeding values (GEBVs) was investigated by considering the GEBVs estimated on offspring as a reference. With AvaII or PstI and ddRADseq with TaqI and PstI, more than 10 K SNPs were detected in common with the HD SNP chip, resulting in an imputation accuracy greater than 0.97. The impact of imputation errors on genomic evaluation of the breeders was reduced, with a Spearman correlation greater than 0.99. Finally, the relative accuracy of GEBVs was equivalent. CONCLUSIONS: RADseq approaches can be interesting alternatives to low-density SNP chips for genomic selection. With more than 10 K SNPs in common with the SNPs of the HD SNP chip, good imputation and genomic evaluation results can be obtained. However, with real data, heterogeneity between individuals with missing data must be considered.

The pigments in eggshell with different colour and the pigment regulatory gene expression in corresponding chicken's shell gland.

Eggshell colour is the unique appearance and economically valuable trait of eggs, whereas the colour is often short of uniformity, especially in the blue-shelled breeds, hence, their pigment differences and molecular mechanism need clarity. To investigate the relationship between the pigment content of eggshells and related gene expression in the eggshell glands of chickens, four subtypes of blue-shelled eggs ('Olive', 'Green', 'Blue', and 'Light') from the same blue-eggshell chicken line were selected; Hy-Line 'White' and 'Brown'-shelled eggs were used as control groups. The L*, a*, b* values, and protoporphyrin-IX and biliverdin contents in each group of eggshells were measured. In addition, the shell glands of the corresponding hens were collected to detect SLCO1B3 genotype and mRNA expression, and ABCG2 and HMOX1 transcription and protein expression. Eggshell colour L* values were negatively correlated with protoporphyrin-IX, b* values were positively correlated with total pigment content (P < 0.001), and a* values were positively correlated with protoporphyrin-IX (P < 0.001) but negatively with biliverdin. Moreover, all four blue-eggshell subtypes were SLCO1B3 homozygous, with SLCO1B3 mRNA expression in shell glands being significantly higher than in the White and Brown groups. ABCG2 and HMOX1 mRNA expression were highest in the Brown and Green groups, respectively (P < 0.05), and were positively correlated with protoporphyrin-IX (P < 0.001) and biliverdin contents in eggshells, respectively. Western blot and immunohistochemical results demonstrated that the Brown group had the highest ABCG2 expression (P < 0.05), followed by the Green and Olive groups. HMOX1 protein expression was higher in the Olive and Green groups (P < 0.05), and lowest in the White group. This study suggests that ABCG2 and HMOX1 have important regulatory roles in the production and transport of protoporphyrin-IX and biliverdin in blue-shelled chicken eggs, respectively.

Genetic and microbiome analysis of feed efficiency in laying hens.

Improving feed efficiency is an important target for poultry breeding. Feed efficiency is affected by host genetics and the gut microbiota, but many of the mechanisms remain elusive in laying hens, especially in the late laying period. In this study, we measured feed intake, body weight, and egg mass of 714 hens from a pedigreed line from 69 to 72 wk of age and calculated the residual feed intake (RFI) and feed conversion ratio (FCR). In addition, fecal samples were also collected for 16S ribosomal RNA gene sequencing (V4 region). Genetic analysis was then conducted in DMU packages by using AI-REML with animal model. Moderate heritability estimates for FCR (h2 = 0.31) and RFI (h2 = 0.52) were observed, suggesting that proper selection programs can directly improve feed efficiency. Genetically, RFI was less correlated with body weight and egg mass than that of FCR. The phenotypic variance explained by gut microbial variance is defined as the microbiability (m2). The microbiability estimates for FCR (m2 = 0.03) and RFI (m2 = 0.16) suggested the gut microbiota was also involved in the regulation of feed efficiency. In addition, our results showed that the effect of host genetics on fecal microbiota was minor in three aspects: 1) microbial diversity indexes had low heritability estimates, and genera with heritability estimates more than 0.1 accounted for only 1.07% of the tested fecal microbiota; 2) the genetic relationship correlations between host genetics and different microbial distance were very weak, ranging from -0.0057 to -0.0003; 3) the microbial distance between different kinships showed no significant difference. Since the RFI has the highest microbiability, we further screened out three genera, including Anaerosporobacter, Candidatus Stoquefichus, and Fournierella, which were negatively correlated with RFI and played positive roles in improving the feed efficiency. These findings contribute to a great understanding of the genetic background and microbial influences on feed efficiency.

Isolation and characterization of avian leukosis virus subgroup J associated with hemangioma and myelocytoma in layer chickens in China.

A strain of avian leukosis virus (ALV) belonging to a new envelope subgroup J (ALV-J) emerged in 1988 as a new subgroup of ALV and spread rapidly throughout the world. Due to the infection and spread of ALV-J, the global poultry industry experienced a significant loss. Although the disease had been prevented and controlled effectively by culling domestic chickens in the infected zone, a few field cases of ALV-J infection were reported in China in recent years. This study was conducted to characterize the genome and analyze the lesions and histopathology of the ALV-J strain named HB2020, which was isolated from layer chickens in Hubei Province, China. The full-length proviral genome sequence analysis of ALV-J HB2020 revealed that it was a recombinant strain of ev-1 and HPRS-103 in the gag gene in comparison to ALV-J prototype HPRS-103. In the 3'-untranslated region (3'UTR) of the nucleotide sequence, there were found 205-base pairs (bp) deletion, of which 175 were detected in the redundant transmembrane (rTM) region. Besides, the surface glycoprotein gene gp85 had five mutations in a conservative site, whereas the transmembrane protein gene gp37 was relatively conserved. The animal experiments conducted later on this strain have shown that HB2020 can cause various neoplastic lesions in chickens, including enlarged livers with hemangiomas and spleens with white nodules. Additionally, as the exposure time increased, the number of tumor cells that resembled myelocytes in the blood smears of infected chickens gradually increased. These results indicated that HB2020 on recombination with ALV subgroup E (ALV-E) and ALV-J could induce severe hemangiomas and myelocytomas. This inference might provide a molecular basis for further research about the pathogenicity of ALV and emphasize the need for control and prevention of avian leukosis.

Determinants of Newcastle disease in commercial layer chicken farms in two districts of Bangladesh: A case-control study.

Newcastle disease (ND) is a real threat for commercial layer chicken farms in Bangladesh. However, only few studies have focused on exploring the epidemiology of this disease. A case-control study was conducted to identify determinants of Newcastle disease in commercial layer chicken farms in Kishoreganj and Gazipur district of Bangladesh between September 2019 and February 2020. Farms with birds diagnosed as ND positive based on clinical history, clinical signs and postmortem findings were considered as case and farms that did not have such ND positive chickens were the control for this study. Farmers of 56 case farms and 56 control farms were interviewed face to face using a structured questionnaire. The association between Farms' ND status and determinants was assessed by multivariable logistic regression with backward elimination. In the final model, six variables were found to be associated with the risk for ND outbreak: age of the farmers (Odds Ratio [OR] 0.94; 95% Confidence Interval [CI] 0.87-0.99), distance from the nearest poultry farms (OR = 3.23, 95% CI 1.27-8.39), number of houses in the farms (OR = 3.06, 95% CI 1.06-8.83), surrounding environments (OR = 5.27, 95% CI 1.96-14.20), rearing different aged bird together (OR = 4.76, 95% CI 1.25-18.19), and no isolation of sick birds (OR = 2.85, 95% CI 1.07-7.55). Alteration of these determinants should reduce the ND burden in commercial layer chicken farms.

Genome-wide association studies for egg quality traits in White Leghorn layers using low-pass sequencing and SNP chip data.

Low-pass sequencing data have been proposed as an alternative to single nucleotide polymorphism (SNP) chips in genome-wide association studies (GWAS) of several species. However, it has not been used in layer chickens yet. This study aims at comparing the GWAS results of White Leghorn chickens using low-pass sequencing data (1×) and 54 k SNP chip data. Ten commercially relevant egg quality traits including albumen height, shell strength, shell colour, egg weight and yolk weight collected from up to 1,420 White Leghorn chickens were analysed. The results showed that the genomic heritability estimates based on low-pass sequencing data were higher than those based on SNP chip data. Although two GWAS analyses showed similar overall landscape for most traits, low-pass sequencing captured some significant SNPs that were not on the SNP chip. In GWAS analysis using 54 k SNP chip data, after including more individuals (up to 5,700), additional significant SNPs not detected by low-pass sequencing data were found. In conclusion, GWAS using low-pass sequencing data showed similar results to those with SNP chip data and may require much larger sample sizes to show measurable advantages.

Efficacy of dietary supplementary probiotics as substitutes for antibiotic growth promoters during the starter period on growth performances, carcass traits, and immune organs of male layer chicken.

Background and Aim: With the increased concerns about global protein supply, chicken meat, especially from male layer chicken, constitutes an alternative in terms of quality and carcass traits. Probiotics have been proposed for replacing antibiotic growth promoters (AGPs), which have been prohibited as poultry supplement feeds. The present study aimed to determine the efficacy of dietary supplementary probiotics during the starter period on growth performances, carcass traits, and immune organs of male layer chicken. Materials and Methods: In this study, one hundred and eighty 1-day-old male chicks from the strain ISA brown were used. They were divided into six groups according to the feed: 100% basal feed (T0), basal feed+2.5 g AGP/kg feed (T1), basal feed+probiotics 1 mL/kg feed (T2), basal feed+probiotics 3 mL/kg feed (T3), basal feed+probiotics 4 mL/kg feed (T4), and basal feed+probiotics 5 mL/kg feed (T5). Probiotics (Lactobacillus acidophilus, Lactobacillus plantarum, and Bifidobacterium spp.) were given at a concentration of 1.2×109 colony-forming unit/mL. Virginiamycin was used as AGP. ISA brown layer chicken was treated for 21 days. Growth performances (body weight, feed consumption, and feed conversion ratio [FCR]), carcass traits (weight at slaughter, weight of the carcass, breast muscles, liver, lungs, kidneys, and heart), immune organs (spleen, thymus, and bursa of Fabricius), and non-edible organs (head, legs, and wings) were analyzed. Results: Probiotic supplementation at 4 and 5 mL/kg feed (T4 and T5) during the starter phase improved the body weight, FCR, and feed consumption. The weight at slaughter, weight of the carcass, breast muscles, and liver from the T4 and T5 groups were significantly greater than those in the other treatment groups. In addition, the weight of the heart, lungs, and kidneys was increased in the T1, T2, T3, T4, and T5 groups compared with that measured in the T0 group. Furthermore, there were significant differences regarding the immune organs between the T0 and the other treatment groups. The weight of the head, legs, and wings was also greater in the probiotic and AGP supplementation groups (T1, T2, T3, T4, and T5) than that in the basal feed group (T0). Conclusion: Probiotic (L. acidophilus, L. plantarum, and Bifidobacterium spp.) supplementation at 4 and 5 mL/kg feed during the starter period can be used to improve the growth, carcass traits, and weight of immune organs in male layer chicken.

Chicken hepatomegaly and splenomegaly associated with novel subgroup J avian leukosis virus infection.

BACKGROUND: Subgroup J avian leukosis virus (ALV-J) is an oncovirus which can induce multiple types of tumors in chicken. In this report, we found novel ALV-J infection is closely associated with serious hepatomegaly and splenomegaly in chicken. CASE PRESENTATION: The layer chickens from six flocks in Jiangsu province, China, showed serious hemoperitoneum, hepatomegaly and splenomegaly. Histopathological results indicated focal lymphocytic infiltration, cell edema and congestion in the liver, atrophy and depletion of lymphocyte in the spleen. Tumor cells were not detected in all the organs. avian hepatitis E virus (aHEV), which is thought to be the cause of a very similar disease, big liver and spleen disease (BLS), was not detected. Other viruses causing tumors or liver damage including Marek's disease virus (MDV), reticuloendotheliosis virus (REV), fowl adenovirus (FAdV) and chicken infectious anemia virus (CIAV) were also proved negative by either PCR or RT-PCR. However, we did detect ALV-J in those chickens using PCR. Only novel ALV-J strains were efficiently isolated from these chicken livers. CONCLUSIONS: This is the first report that chicken hepatomegaly and splenomegaly disease was closely associated with novel ALV-J, highlighting the importance of ALV-J eradication program in China.

Route of infectious bronchitis virus vaccination determines the type and magnitude of immune responses in table egg laying hens.

Chicken immune responses to infectious bronchitis virus (IBV) vaccination can depend on route of administration, vaccine strain and bird age. Typically for layer chickens, IBV vaccinations are administered by spray in the hatchery at day-old and boosted at intervals with live vaccines via drinking water (DW). Knowledge of live attenuated IBV vaccine virus kinetics and the immune response in egg-laying hens is exceptionally limited. Here, we demonstrated dissemination of vaccine viruses and differences in hen innate, mucosal, cellular and humoral immune responses following vaccination with Massachusetts or 793B strains, administered by DW or oculonasal (ON) routes. Detection of IBV in the Mass-vaccinated groups was greater during early time-points, however, 793B was detected more frequently at later timepoints. Viral RNA loads in the Harderian gland and turbinate tissues were significantly higher for ON-Mass compared to all other vaccinated groups. Lachrymal fluid IgY levels were significantly greater than the control at 14 days post-vaccination (dpv) for both vaccine serotypes, and IgA mRNA levels were significantly greater in ON-vaccinated groups compared to DW-vaccinated groups, demonstrating robust mucosal immune responses. Cell mediated immune gene transcripts (CD8-α and CD8-ß) were up-regulated in turbinate and trachea tissues. For both vaccines, dissemination and vaccine virus clearance was slower when given by DW compared to the ON route. For ON administration, both vaccines induced comparable levels of mucosal immunity. The Mass vaccine induced cellular immunity to similar levels regardless of vaccination method. When given either by ON or DW, 793B vaccination induced significantly higher levels of humoral immunity.

Interaction of Central Glutamatergic and Histaminergic Systems on Food Intake Regulation in Layer Chickens.

This study purposed to discover the connection between the central glutamatergic and histaminergic systems on feeding behavior in layer chickens. In the first experiment, chicks obtained intracerebroventricular (ICV) injections of saline (control solution), α-FMH (250 nmol), glutamate (300 nmol), and α-FMH + glutamate. Experiments 2-6 were comparable to the first experiment, apart from the birds being injected with chlorpheniramine (histamine H1 receptor antagonist, 300 nmol), famotidine (histamine H2 receptor antagonist, 82 nmol), and thioperamide (histamine H3 receptor antagonist, 300 nmol) instead of α-FMH. In Experiment five, experimental groups were divided into (A) control solution, (B) MK-801 (N-methyl-D-aspartate receptor antagonist, 15 nmol), (C) histamine (300 nmol) and (D) MK-801 + histamine. Experiments 6-10 and Experiment five were similar apart from the ICV injections of CNQX (AMPA receptor antagonist, 360 nm), UBP-302 (Kainate receptor antagonist, 390 nm), AIDA (mGluR1 antagonist, 2 nmol), LY341495 (mGluR2 antagonist, 150 nmol), and UBP1112 (mGluR3 antagonist, 2 nmol) given instead of MK-801. Afterward, cumulative food intake was recorded at30, 60, and 120 minutes after the injection process. According to the results, ICV injection of glutamate considerably reduced food intake (p<0.05). Co-injection of α-FMH + glutamate and/or chlorpheniramine + glutamate reduced the hypophagic influence of glutamate (p<0.05), whereas thioperamide + glutamate augmented glutamate-induced hypophagia in neonatal chicks (p<0.05). Co-injection of MK-801 + histamine or UBP-302 + histamine reduced the hypophagic influence of the histamine (p<0.05), whereas LY341495 + histamine augmented the hypophagic influence of the histamine (p<0.05). Given the results, it is suggested that the effect of the connection between these systems on the process of food intake regulation is mediated by H1 and H3 histamines as well as NMDA, Kainate, and mGluR2 glutamate receptors in neonatal layer chickens.

Genomic Characterization of Fluoroquinolone-Resistant Thermophilic Campylobacter Strains Isolated from Layer Chicken Feces in Gangneung, South Korea by Whole-Genome Sequencing.

Thermophilic Campylobacter species of poultry origin have been associated with up to 80% of human campylobacteriosis cases. Layer chickens have received less attention as possible reservoirs of Campylobacter species. Initially, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of two archived Campylobacter isolates (Campylobacter jejuni strain 200605 and Campylobacter coli strain 200606) from layer chickens to five antimicrobials (ciprofloxacin, nalidixic acid, erythromycin, tetracycline, and gentamicin) were determined using broth microdilution while the presence of selected antimicrobial resistance genes was performed by polymerase chain reaction (PCR) using specific primers. Whole-genome sequencing (WGS) was performed by the Illumina HiSeq X platform. The analysis involved antimicrobial resistance genes, virulome, multilocus sequence typing (MLST), and phylogeny. Both isolates were phenotypically resistant to ciprofloxacin (MIC: 32 vs. 32 µg/mL), nalidixic acid (MIC: 128 vs. 64 µg/mL), and tetracycline (MIC: 64 vs. 64 µg/mL), but sensitive to erythromycin (MIC: 1 vs. 2 µg/mL) and gentamicin (MIC: 0.25 vs. 1 µg/mL) for C. jejuni strain 200605 and C. coli strain 200606, respectively. WGS confirmed C257T mutation in the gyrA gene and the presence of cmeABC complex conferring resistance to FQs in both strains. Both strains also exhibited tet(O) genes associated with tetracycline resistance. Various virulence genes associated with motility, chemotaxis, and capsule formation were found in both isolates. However, the analysis of virulence genes showed that C. jejuni strain 200605 is more virulent than C. coli strain 200606. The MLST showed that C. jejuni strain 200605 belongs to sequence type ST-5229 while C. coli strain 200606 belongs to ST-5935, and both STs are less common. The phylogenetic analysis clustered C. jejuni strain 200605 along with other strains reported in Korea (CP028933 from chicken and CP014344 from human) while C. coli strain 200606 formed a separate cluster with C. coli (CP007181) from turkey. The WGS confirmed FQ-resistance in both strains and showed potential virulence of both strains. Further studies are recommended to understand the reasons behind the regional distribution (Korea, China, and Vietnam) of such rare STs.

Interconnection between Adrenergic and Dopaminergic Systems in Feeding Behavior in Neonatal Chicks.

Central dopaminergic (DAergic) and adrenergic systems have a prominent role in appetite regulation; however, their interaction(s) have not been studied in neonatal layer chickens.Therefore, the current study aimed to determine the interaction of central DAergic and noradrenergic systems in food intake regulation in neonatal layer chickens. In the first experiment, chickens received the intracerebroventricular (ICV) injection of a control solution, prazosin (i.e., &alpha;1 adrenergic receptor antagonist; 10 nmol), dopamine (DA; 40 nmol), and prazosin plus DA. The second to fifth experiments were similar to the first experiment except that the birds were injected with yohimbine (i.e., &alpha;2 receptor antagonist; 13 nmol), metoprolol (i.e., &beta;1 adrenergic receptor antagonist; 24 nmol), ICI 118,551 (i.e., &beta;2 adrenergic receptor antagonist; 5 nmol), and SR59230R (i.e., &beta;3 adrenergic receptor antagonist; 20 nmol) instead of prazosin. In the sixth experiment, the chickens received ICV injection with the control solution and noradrenaline (NA; 75, 150, and 300 nmol). In the seventh experiment, the birds were injected with the control solution, SCH23390 (i.e., D1 DAergic receptor antagonist; 5 nmol), NA (300 nmol), and SCH23390 plus NA In the eighth experiment, the control solution, AMI-193 (i.e., D2 DAergic receptor antagonist; 5 nmol), NA (300 nmol), and AMI-193 plus NA were injected. Then, cumulative food intake was recorded at 30, 60, and 120 min after the injection. According to the obtained results, the ICV injection of DA (40 nmol) significantly decreased food intake in comparison to that reported for the control group (p &lt;0.05). The co-injection of yohimbine plus DA significantly amplified DA-induced hypophagia in the neonatal chickens (p &lt;0.05). In addition, the co-administration of ICI 118,551 plus DA significantly inhibited the hypophagic effect of DA in the neonatal chickens (p &lt;0.05). Furthermore, NA (75, 150, and 300 nmol) significantly reduced food intake in a dose-dependent manner (p &lt;0.05). The co-injection of SCH23390 plus NA decreased the hypophagic effect of NA in the neonatal chickens, compared to that reported for the control group (p &lt;0.05). The co-injection of AMI-193 plus NA diminished NA-induced hypophagia, compared to that reported for the control group (p &lt;0.05). The aforementioned results suggested that there is an interconnection between central DAergic and noradrenergic systems through &alpha;2/&beta;2 adrenergic and D1/D2 DAergic receptors in food intake regulation in neonatal chicks.

Understanding the effects of intramuscular injection and feed withdrawal on Salmonella Typhimurium shedding and gut microbiota in pullets.

BACKGROUND: Gut microbiota plays a key role in health, immunity, digestion, and production in layers. Factors such as environment, diet, diseases, stress, and flock management significantly affect gut microbiota; however, it is not known how potential stressors such as intramuscular injections or feed withdrawal alter the composition of gut microbiota that result in increased the shedding level of foodborne pathogens. In the current study, the effects of intramuscular corticosterone injection and feed withdrawal were evaluated to understand their role in Salmonella Typhimurium shedding and changes in the composition of gut microbiota in layers. RESULTS: Salmonella shedding was observed for 8 weeks post-infection. There was a significant increase in Salmonella Typhimurium count after intramuscular injection and feed withdrawal. The Salmonella infected and the negative control groups showed significant differences in the abundance of different genera in gut microbiota at week 1 and up to week 7 post infection. The infected group showed a significant reduction in alpha diversity of gut microbiota. Firmicutes reduced significantly (P < 0.05) after intramuscular injection, while the feed withdrawal groups did not cause any significant changes in Proteobacteria-Firmicutes ratio. Furthermore, intramuscular injection resulted in a significant change in alpha diversity of gut microbiota. CONCLUSIONS: Exposure of chicks to relatively low dose of Salmonella Typhimurium can lead to persistent shedding in pullets. The Salmonella Typhimurium infection disrupted the gut microbiota composition immediately after infection. The potential stress of intramuscular injection and feed withdrawal significantly increased the Salmonella Typhimurium count in faeces. The intramuscular injection also resulted in a significant alteration of the Proteobacteria-Firmicutes ratio, which could increase the risk of dysbiosis.

Impact of the Central Histaminergic and Melanocortin Systems on Leptin-Induced Hypophagia in Neonatal Layer Chicken.

The present study aimed to assess the probable impact of the central histaminergic and melanocortin systems on leptin-induced hypophagia in neonatal layer chickens. In experiment 1, the chickens received intracerebroventricular (ICV) injections of the control solution, 250 nmol of α-FMH, 10 µg of leptin, and α-FMH+leptin. Experimental groups 2-8 were injected the same as experiment 1. Nonetheless, the chickens in experiments 2-8 received ICV injections of 300 nmol of chlorpheniramine (H1 receptor antagonist), 82 nmol of famotidine (H2 receptor antagonist), 300 nmol of thioperamide (H3 receptor antagonist), 0.5 nmol of SHU9119 (M3/M4 receptors antagonist), 0.5 nmol of MCL0020 (M4 receptor antagonist), 30 µg of astressin-B (CRF1/ CRF2 receptors antagonist), and 30 µg of astressin2-B (CRF2 receptor antagonist), instead of α-FMH, respectively. Food was provided for the birds immediately following the injection, and 30, 60, and 120 min after the injection, cumulative food intake (g) was measured. The findings pointed out that the ICV injection of leptin diminished food intake in neonatal chickens (P<0.05). The co-administration of M3/M4 receptor antagonist+leptin significantly decreased the hypophagic effect of leptin (P<0.05). A significant decrease was also detected in the hypophagic effect of leptin following the co-administration of the M4 receptor antagonist and leptin (P<0.05). Moreover, the co-injection of the antagonists of CRF1/CRF2 receptors and leptin significantly mitigated the hypophagic effect of leptin (P<0.05). The co-injection of CRF2 receptor antagonist and leptin led to a decrease in the hypophagic effect of leptin. As evidenced by the results of the current study the hypophagic effect of leptin is mediated by the receptors of H1, H3, M3/M4, and CRF1/CRF2 in neonatal layer chicken.