Analysis of the effects of ß-mannanase on immune function and intestinal flora in broilers fed the low energy diet based on 16S rRNA sequencing and metagenomic sequencing.

As an enzyme, ß-mannanase (BM) can be widely used as feed additive to improve the growth performance of animals. This experiment aimed to determine the effect of the addition of BM to low-energy diet on the immune function and intestinal microflora of broiler chickens. In this study, 384 one-day-old Arbor Acres broilers were randomly divided into 3 groups (8 replicates per group): positive control (PC, received a corn-soybean meal basal diet), negative control (NC, received a low-energy diet with Metabolizable Energy (ME) reduced by 50 kcal/kg) and NC + BM group (NC birds + 100 mg/kg BM). All birds were raised for 42 d. The results showed that BM mitigated the damage of immune function in peripheral blood of broilers caused by the decrease of dietary energy level by increasing the Concanavalin A (Con A) index of stimulation (SI) and macrophages phagocytic activity in the peripheral blood of broilers at 42 d (P < 0.05). The analysis of cecum flora showed that the low-energy diet significantly reduced the observed_species index (P < 0.01), Chao1 index and ACE index (P < 0.05), which reduced the abundance and evenness of species in the cecum of broilers at 21 d. It also significantly reduced the relative abundance of Candidatus_Arthromitus and significantly increased the relative abundance of Pseudomonas in the cecum of broilers at 21 d, while also significantly increasing the relative abundance of Monoglobus at 42 d. BM significantly increased the relative abundance of Lachnospiraceae_UCG-001 and Lachnospiraceae_bacterium_615 in the cecum of broilers at 21 d. In addition, BM inhibited microbial Fatty acid degradation by decreasing the activity of glutaryl-CoA dehydrogenase. Collectively, BM could improve intestinal health by enhancing the immune function of broilers, promoting the proliferation of beneficial bacteria and reducing the number of harmful bacteria, regulating intestinal flora, thereby alleviating the adverse effects of lower dietary energy levels.

The accessible chromatin landscape of lipopolysaccharide-induced systemic inflammatory response identifying epigenome signatures and transcription regulatory networks in chickens.

Lipopolysaccharide (LPS) can induce systemic inflammatory response (SIR) in animals. Understanding the regulatory mechanism of SIR and therapies to ensure healthy growth is urgently needed. Chromatin remodeling plays a crucial role in the expression of genes involved in immune diseases. In the present study, the ATAC-seq analysis revealed 3491 differential open chromatin sites in the spleen of chicks with SIR induced by LPS challenge, and we presented the motifs on these sites and the associated transcription factors. The regulatory network was presented by combining the differential open chromatin data with the mRNAs and exploded cytokines. Interestingly, the LPS challenge could regulate the mRNA expression of 202 genes through chromatin reprogramming, including critical genes such as TLE1 and JUN, which regulate signaling pathways such as I-κB kinase/NF-κB, Toll-like receptor, and downstream cytokine genes. Furthermore, dietary daidzein could inhibit DNA topoisomerase II, which reprograms the spatial conformation of chromatin in the inflammatory response and attenuates SIR. In conclusion, we successfully identified key genes directly regulated by chromatin reprogramming in SIR and demonstrated the chromatin epigenome signatures and transcriptional regulatory network, which provides an important reference for further research on avian epigenetics. There is great potential for alleviating SIR using dietary daidzein.

Probiotic cocktails accelerate baicalin metabolism in the ileum to modulate intestinal health in broiler chickens.

BACKGROUND: Baicalin and probiotic cocktails are promising feed additives with broad application prospects. While probiotic cocktails are known to enhance intestinal health, the potential synergistic impact of combining baicalin with probiotic cocktails on the gut health of broiler chickens remains largely unexplored. Therefore, this study aims to investigate the influence of the combined administration of baicalin and probiotic cocktails on the composition of ileal and cecal microbiota in broiler chickens to elucidate the underlying mechanisms responsible for the health-promoting effects. RESULTS: A total of 320 1-day-old male Arbor Acres broilers were divided into 4 groups, each with 8 replicates of 10 chicks per replicate. Over a period of 42 d, the birds were fed a basal diet or the same diet supplemented with 37.5 g/t baicalin (BC), 1,000 g/t probiotic cocktails (PC), or a combination of both BC (37.5 g/t) and PC (1,000 g/t). The results demonstrated that BC + PC exhibited positive synergistic effects, enhancing intestinal morphology, immune function, and barrier function. This was evidenced by increased VH/CD ratio, sIgA levels, and upregulated expression of occludin and claudin-1 (P < 0.05). 16S rRNA analysis indicated that PC potentiated the effects of BC, particularly in the ileum, where BC + PC significantly increased the α-diversity of the ileal microbiota, altered its ß-diversity, and increased the relative abundance of Flavonifractor (P < 0.05), a flavonoid-metabolizing bacterium. Furthermore, Flavonifractor positively correlated with chicken ileum crypt depth (P < 0.05). While BC + PC had a limited effect on cecal microbiota structure, the PC group had a very similar microbial composition to BC + PC, suggesting that the effect of PC at the distal end of the gut overshadowed those of BC. CONCLUSIONS: We demonstrated the synergistic enhancement of gut health regulation in broiler chickens by combining baicalin and probiotic cocktails. Probiotic cocktails enhanced the effects of baicalin and accelerated its metabolism in the ileum, thereby influencing the ileal microbiota structure. This study elucidates the interaction mechanism between probiotic cocktails and plant extract additives within the host microbiota. These findings provide compelling evidence for the future development of feed additive combinations.

Mannanase improves the growth performance of broilers by alleviating inflammation of the intestinal epithelium and improving intestinal microbiota.

This experiment aimed to discuss and reveal the effect and mechanism of mannanase on intestinal inflammation in broilers triggered by a soybean meal diet. In this experiment, 384 Arbor Acres broilers at 1 d old were randomly divided into 3 treatment groups. The broilers were fed a corn-soybean meal basal diet, a low-energy diet (metabolizable energy reduced by 50 kcal/kg), and a low-energy diet supplemented with 100 mg/kg mannanase for 42 d. The low-energy diet increased feed conversion ratio from 0 to 42 d, reduced ileal villus height and villus height-to-crypt depth ratio and upregulated the expression of nuclear factor kappa B (NF-κB) in the ileum (P < 0.05). It also reduced cecal short-chain fatty acids (SCFA), such as acetic acid (P < 0.05). Compared with low-energy diets, the addition of mannanase increased body weight at 42 d, promoted the digestibility of nutrients, and maintained the morphology and integrity of the intestinal epithelium of broilers (P < 0.05). In addition, mannanase upregulated the expression of claudin-1 (CLDN1) and zonula occludens-1 (ZO-1) in the jejunum at 21 d, downregulated the expression of ileal NF-κB, and increased the content of isobutyric acid in the cecum of broilers (P < 0.05). The results for the ileal microbiota showed that a low-energy diet led to a decrease in the relative abundance of Lactobacillus reuteri in the ileum of broilers. The addition of mannanase increased the relative abundance of Lactobacillus-KC45b and Lactobacillus johnsonii in broilers. Furthermore, a low-energy diet reduced the relative abundance of Butyricicoccus in the intestine of broilers and inhibited oxidative phosphorylation and phosphoinositol metabolism. Mannanase increased the relative abundance of Odoribacter, promoted energy metabolism and N-glycan biosynthesis, and increased the activities of GH3 and GH18. It is concluded that mannanase could improve the growth performance of broilers by reducing the expression of NF-κB in the ileum, increasing the production of SCFA in the cecum, suppressing intestinal inflammation, balancing the intestinal microbiota, reducing damage to the intestinal barrier, and improving the efficiency of nutrient utilization to alleviate the adverse effects caused by the decrease in dietary energy level.

The ionome and proteome landscape of aging in laying hens and relation to egg white quality.

The ionome is essential for maintaining body function and health status by participating in diverse key biological processes. Nevertheless, the distribution and utilization of ionome among different organs and how aging impacts the ionome leading to a decline in egg white quality remain unknown. Thus, we used inductively coupled plasma mass spectrometry (ICP-MS) to analyze 35 elements and their isotopic contents in eight organs of laying hens at 35, 72, and 100 weeks. Moreover, the magnum proteome, amino acids in egg white, and egg white quality were analyzed in laying hens at three different ages using 4D proteomics techniques, an amino acid analyzer, and an egg quality analyzer. Across the organs, we identified varying distribution patterns among macroelements (Mg24, Ca43/44, K39, and P31), transition metals (Zn64/66, Cu63/65, Fe56/57, and Mn55), and toxic elements (Pb208, Ba137, and Sr86). We observed an organ-specific aging pattern characterized by the accumulation of toxic elements (Pb208, Ba137, and Sr86) and calcification in the small intestine. Additionally, a decrease in the utilization of essential trace elements selenium (Se78/82) and manganese (Mn55) was noted in the oviduct. By analyzing ionome in tandem with egg quality, egg white amino acids, and proteome, we unveiled that the reduction of selenium and manganese concentrations in the magnum during the aging process affected amino acid metabolism, particularly tryptophan metabolism, thereby inhibiting the amino acid synthesis in the magnum. Furthermore, it accelerated the senescence of magnum cells through necroptosis activation, leading to a decline in the albumen secretion function of the magnum and subsequently reducing egg white quality. Overall, this study provides insights into the evolution of 35 elements and their isotopes across 8 organs of laying hens with age. It also reveals the elemental composition, interactions, and utilization patterns of these organs, as well as their correlation with egg white quality. The present study highlights the significance of ionome and offers a comprehensive perspective on the selection of ionome for regulating the aging of laying hens.

Effects of Dietary Supplemental Chlorogenic Acid and Baicalin on the Growth Performance and Immunity of Broilers Challenged with Lipopolysaccharide.

The objective of this study was to investigate the effects of dietary supplemental chlorogenic acid and baicalin (CAB) on the growth performance and immunity of broilers challenged with lipopolysaccharide (LPS). This study was designed as a factorial arrangement of 2 dietary CAB treatments × 2 LPS treatments. Birds challenged with or without LPS were fed with a basic diet (CON) and (LPS), the level of CAB diet containing 500 mg/kg CAB(CAB) and (CAB + LPS). The feeding trial lasted for 42 days. Results showed that there was a negative effect on average daily weight gain (ADG) and average body weight of broilers during the animal trial with LPS challenge. The levels of diamine oxidase (DAO), lysozyme (LYZ), immunoglobulin G (IgG), and IgA in the serum, the contents of IL-1ß and TNF-α in the spleen were elevated with LPS treated. Additionally, LPS treatment tended to reduce the jejunal villi height (VH) and total superoxide dismutase (T-SOD) in the serum. Dietary supplemental 500 mg/kg CAB increased the body weight and ADG and improved the feed conversion ratio (FCR) during the trial period. In addition, dietary 500 mg/kg CAB elevated the ratio of VH to crypt depth in the jejunum and reduced the content of protein carbonyl. Beyond that, the levels of IgG and IgA in the serum and transforming growth factor (TGF-ß) in the spleen were up-regulated with 500 mg/kg CAB supplementation. In conclusion, dietary CAB was beneficial for growth performance and immunity of broilers challenged with lipopolysaccharide.

Supplementing Genistein for Breeder Hens Alters the Growth Performance and Intestinal Health of Offspring.

Recent research revealed that dietary genistein supplementation for breeder hens can improve the immune function of offspring chicks. However, it remains unknown whether this maternal effect could improve the intestinal health of offspring. This study was conducted to explore the mechanism involved in the maternal effect of genistein on the intestinal mucosa and microbial homeostasis of chicken offspring. A total of 120 Qiling breeder hens were fed a basal diet, a 20 mg/kg genistein-supplemented diet, or a 40 mg/kg genistein-supplemented diet for 4 weeks before collecting their eggs. After hatching, 180 male offspring (60 chickens from each group) were randomly selected and divided into three groups: (1) the offspring of hens fed a basal diet (CON); (2) the offspring of hens fed a low-dose genistein-supplemented diet (LGE); (3) the offspring of hens fed a high-dose genistein-supplemented diet (HGE). At 17 d, 72 male offspring (48 chickens from CON and 24 chickens from LGE) were divided into three groups: (1) the offspring of hens fed a basal diet (CON); (2) the CON group challenged with LPS (LPS); (3) the LGE group challenged with LPS (LPS + LGE). The results showed that maternal genistein supplementation increased the birth weight and serum level of total protein (TP), followed by improved intestinal villus morphology. Continuously, the maternal effect on the body weight of chicks lasted until 21 d. Additionally, it was observed that maternal genistein supplementation exhibited protective effects against LPS-induced morphological damage and intestinal mucosal barrier dysfunction by upregulating the expression of tight junction proteins, specifically ZO-1, Claudin1, E-cadherin, and Occludin, at 21 d. Using 16S rRNA gene sequencing, we demonstrated that maternal supplementation of genistein has the potential to facilitate the maturation of newly hatched chicken offspring by enhancing the abundance of Escherichia coli. Additionally, maternal genistein supplementation can effectively reduce the abundance of Gammaproteobacteria, thus mitigating the risk of bacterial diversity impairment of LPS. In light of these findings, maternal genistein supplementation holds promise as a potential strategy for ameliorating intestinal mucosal damage and modulating the microbiome in chicken offspring.

Effects of Tannic Acid Supplementation on the Intestinal Health, Immunity, and Antioxidant Function of Broilers Challenged with Necrotic Enteritis.

Clostridium perfringens causes necrotic enteritis (NE) after proliferation in the intestine of poultry, resulting in considerable losses to the poultry industry. This study aimed to investigate the impact of tannic acid on the antioxidant, immunity, and gut health of broilers with NE. In the experiment, 630 one-day-old Cobb500 male chicks were randomly divided into six treatment groups, with seven replicate cages and with fifteen birds in each cage. The treatment groups were as follows: control group (NC), challenged group (PC), and challenged NE chickens treated with 250, 500, 750, and 1000 mg/kg tannic acid (PTA1, PTA2, PTA3, and PTA4, respectively). To induce NE, coccidia vaccine and Clostridium perfringens were administered on day 19 and days 22-28, respectively. Indexes related to antioxidant, immune, and intestinal health were measured on days 28 and 35. During the infection period, we observed significant increases in fecal water content, D-LA, TNF-α, and malondialdehyde concentrations (p < 0.05). Conversely, significant decreases were noted in chyme pH and in T-AOC, IL-4, and IL-10 concentrations (p < 0.05). The addition of tannic acid exhibited a linear decrease in fecal water content and TNF-α concentration (p < 0.05). Furthermore, tannic acid supplementation resulted in a quadratic curve decrease in D-LA concentration and linear increases in T-AOC, IL-4, and IL-10 (p < 0.05). Cecal microbiological analysis revealed that Ruminococcaceae and Butyricimona were dominant in PTA3. In conclusion, the dietary addition of tannic acid may reduce the negative effects of NE by increasing antioxidant and anti-inflammatory capacity, improving the intestinal barrier, and regulating the intestinal flora.

Effects of tannic acid on the immunity and intestinal health of broiler chickens with necrotic enteritis infection.

BACKGROUND: In broiler chickens, necrotic enteritis (NE) infection can reduce production performance. Tannic acid has shown great potential as a treatment of NE in broilers. However, the appropriate dosage of tannic acid in NE of broilers and the improvement effect on intestinal health are not very clear. In this study, we aimed to investigate the effects of different doses of tannic acid on the production performance, immunity, and intestinal health of broilers by constructing an NE model with C. perfringens infection and determining the appropriate dosage of tannic acid with regard to NE. RESULTS: Challenged birds showed significant reduction in body weight, villus height, and the ratio of villus height to crypt depth (P < 0.05) and increase in the feed consumption gain ratio, intestinal lesion score, and crypt depth (P < 0.05). The infection significantly reduced the relative Bacteroidota and Ligilactobacillus abundance (P < 0.05) and increased the ratio of Firmicutes/Bacteroidota and cecal content of C. perfringens (P < 0.05). Challenged birds fed diets supplemented with tannic acid showed significantly increased mRNA expression of nutrient transport carriers and intestinal barrier genes and growth performance and reduced serum zonulin and endotoxin levels (P < 0.05). Addition of tannic acid to the diet inhibited the inflammatory response by reducing the number of coccidia oocysts in feces and the content of C. perfringens in the cecum. Specifically, tannic acid reduced the serum levels of C reactive protein, myeloperoxidase, and specific IgY and ileal mucosal secretory immunoglobulin A levels in the ileal mucosa compared with those in the NE-infected birds. NE-infected birds fed diets supplemented with tannic acid also showed significantly increased relative Anaerocolumna, Thermoanaerobacterium, and Thermosinus abundance (P < 0.05); their microbial composition and functional predictions were similar to those of the NC group. CONCLUSIONS: Tannic acid in the diet alleviated NE by enhancing the intestinal barrier and absorption function. The recommended dietary tannic acid additive level is 500-750 mg/kg. Our study findings would be useful in reducing related economic losses in the broiler industry.

Quantitative proteomics provides insights into the mechanism of the differences in heat-induced gel properties for egg white proteins with different interior quality during ageing in laying hens.

The purpose of this study was to investigate the mechanism for the differences in heat-induced gel properties of egg white proteins with different interior quality during ageing in laying hens. Quantitative proteomic analysis revealed that the abundance of ovotransferrin, avidin, mucin 5B, and clusterin increased with decreasing Haugh units (HU), leading to the transition from disorder to order in the secondary and tertiary structure of egg white proteins, with the burial of hydrophobic groups and a reduction in the negative charge on the protein surface, rendering the egg white protein solution aggregated. These changes would accelerate the rate of aggregation of egg white proteins during heating, resulting in the loss of orientation of the molecular chains, forming coarse and porous gel structures and poor gel properties. This research provides a new idea for improving the gelling properties of egg whites from lower interior quality during ageing in laying hens.

Construction of low intestinal bacteria model and its effect on laying performance and immune function of laying hens.

The objective of this study was to establish a low-bacteria intestinal model in chickens, and then to investigate the characteristics involving in immune function and intestinal environment of this model. A total of 180 twenty-one-week-old Hy-line gray layers were randomly allocated into 2 treatment groups. Hens were fed with a basic diet (Control), or an antibiotic combination diet (ABS) for 5 weeks. Results showed that the total bacteria in the ileal chyme were significantly dropped after ABS treatment. Compared with the Control group, the genus-level bacteria such as Romboutsia, Enterococcus, and Aeriscardovia were reduced in the ileal chyme of the ABS group (P < 0.05). In addition, the relative abundance of Lactobacillus_delbrueckii, Lactobacillus_aviarius, Lactobacillus_gasseri, and Lactobacillus_agilis in the ileal chyme were also descended (P < 0.05). However, Lactobacillus_coleohominis, Lactobacillus_salivarius, and Lolium_perenne were elevated in the ABS group (P < 0.05). Beyond that, ABS treatment decreased the levels of interleukin-10 (IL-10) and ß-defensin 1 in the serum, as well as the number of goblet cells in the ileal villi (P < 0.05). Additionally, the genes mRNA levels of the ileum such as Mucin2, Toll-like receptors 4 (TLR4), Myeloid differentiation factor 88 (MYD88), NF-κB, IL-1ß, Interferon-gama (IFN-γ), IL-4 and the ratio of IFN-γ to IL-4 were also down-regulated in the ABS group (P < 0.05). In addition, there were no significant changes about egg production rate and egg quality in the ABS group. In conclusion, dietary supplemental antibiotic combination for 5 weeks could establish a low intestinal bacteria model of hens. The establishment of a low intestinal bacteria model did not affect the egg-laying performance, while caused immune suppression in laying hens.

Comparative study of apparent metabolizable energy and net energy values of dephenolized cottonseed proteins for laying hens.

Optimizing the energy utilization of nutrients and ensuring maximum benefits are continuous goals for livestock producers. The net energy (NE) value of feed reflects its nutritional value in the precision feeding system. An experiment was conducted to determine the apparent metabolizable energy (AME) and NE values of 3 types of dephenolized cottonseed protein (DCP) for Hy Line Brown hens aged 42 to 45 weeks using the reference diet substitution method. A reference diet based on corn soybean meal was used to meet the nutritional needs of Hy Line Brown laying hens. To render the crude protein and energy values of the 3 test diets similar, 10.5%, 12%, and 16% of the gross energy yielding ingredients from the reference diet were replaced with DCP 1, DCP 2, and DCP 3, respectively. The birds were fed 4 diets during a 7-d adaptation period. After the dietary adaptation period, 2 birds per replicate from each treatment group were placed in an individual open circuit respiratory calorimetry chamber for a 3-d experimental period. Daily O2 consumption and CO2 production were recorded, and excreta samples were collected. The AME values of DCP 1, DCP 2, and DCP 3 were 3,049.05, 2,820.13, and 2,982.31 kcal/kg of dry matter (DM), respectively. The NE values of DCP 1, DCP 2, DCP 3 were 1,475.77, 1,910.31, and 1,905.37 kcal/kg of DM, respectively, and the NE:AME ratios were 48.40%, 67.74%, and 63.89%, respectively. Our data show that the AME value of DCP does not reflect the nutritional value of the feed. The NE value of DCP with a high ME value was not necessarily high.

Effects of rearing system and antibiotic treatment on immune function, gut microbiota and metabolites of broiler chickens.

BACKGROUND: In China, cage systems with a high space utilization have gradually replaced ground litter systems, but the disease incidence of chickens in cages is higher. Broilers in the ground litter pens may be stimulated by more environmental microbes during the growth process and show strong immune function and status, but knowledge of which microbes and their metabolites play an immunomodulatory role is still limited. This study aimed to explore the differences and correlations in the immune function, gut microbiota and metabolites and the importance of gut microbiota of broilers raised in cages and ground litter pens. METHODS: The experiment involved a 2 × 2 factorial arrangement, with rearing systems (cages or ground litter pens) and antibiotic treatment (with or without broad-spectrum antibiotics in drinking water) as factors. RESULTS: The results showed that, compared with the cage group, the ground litter broilers had stronger nonspecific immune function (Macrophages% and NO in blood), humoral immune function (IgG in blood, LPS stimulation index in ileum) and cellular immune function (T%, Tc%, ConA stimulation index and cytokines in blood). Antibiotic (ABX) treatment significantly reduced nonspecific immune function (Macrophages% and NO in blood, iNOS and Mucin2 mRNA expression in ileum), humoral immune function (IgG in blood and sIgA in ileum) and cellular immune function (T% and cytokines in blood, Th and Tc ratio, TLRs and cytokines mRNA expression in ileum). Furthermore, the ground litter broilers had higher α diversity of microbiota in ileum. The relative abundance of Staphylococcus, Jeotgalicoccus, Jeotgalibaca and Pediococcus in the ileum of ground litter broilers were higher. ABX treatment significantly reduced the α diversity of ileal microbiota, with less Chloroplast and Mitochondria. In addition, the levels of acetic acid, isobutyric acid, kynurenic acid and allolithocholic acid in the ileum of ground litter broilers were higher. Spearman correlation analysis showed that Jeotgalibaca, Pediococcus, acetic acid, kynurenic acid and allolithocholic acid were related to the immune function. CONCLUSIONS: There were more potential pathogens, litter breeding bacteria, short-chain fatty acids, kynurenine, allolithocholic acid and tryptophan metabolites in the ileum of broilers in ground litter pens, which may be the reason for its stronger immune function and status.

Fecal Microbiota Transplantation Reshapes the Physiological Function of the Intestine in Antibiotic-Treated Specific Pathogen-Free Birds.

The topic about the interactions between host and intestinal microbiota has already caught the attention of many scholars. However, there is still a lack of systematic reports on the relationship between the intestinal flora and intestinal physiology of birds. Thus, this study was designed to investigate it. Antibiotic-treated specific pathogen-free (SPF) bird were used to construct an intestinal bacteria-free bird (IBF) model, and then, the differences in intestinal absorption, barrier, immune, antioxidant and metabolic functions between IBF and bacteria-bearing birds were studied. To gain further insight, the whole intestinal flora of bacteria-bearing birds was transplanted into the intestines of IBF birds to study the remodeling effect of fecal microbiota transplantation (FMT) on the intestinal physiology of IBF birds. The results showed that compared with bacteria-bearing birds, IBF birds had a lighter body weight and weaker intestinal absorption, antioxidant, barrier, immune and metabolic functions. Interestingly, FMT contributed to reshaping the abovementioned physiological functions of the intestines of IBF birds. In conclusion, the intestinal flora plays an important role in regulating the physiological functions of the intestine.

Comparison and Correlation Analysis of Immune Function and Gut Microbiota of Broiler Chickens Raised in Double-Layer Cages and Litter Floor Pens.

This study aimed to compare the immune function and gut microbiota between double-layer caged and litter floor pen-raised broiler chickens. Eighty meaty male chicks were selected and divided into cage group and litter floor group, with 20 replicates in each group. The broilers were raised in the same chicken house. The rearing density of the two rearing systems was same. The broilers were sampled on days 13 and 34. The results showed that compared with the cage group, the litter floor broilers had worse growth performance (23.24% increase in feed conversion ratio) in the early stage; better slaughter performance at day 42; stronger peripheral immune function (including higher lysozyme activity, T-cell ratio, Th-cell ratio, Tc-cell ratio, CD4/CD8, IL-10, B-cell ratio, IgG and IgA levels; and spleen immune-related gene expression); and stronger intestinal immune function (including higher ileum CD80, AvBD2, Mucin2, NF-κB, IL-8, IFN-γ/IL-4, and IgA mRNA expression levels and ileal mucosa sIgA levels). Compared with the cage group, the alpha diversity of ileum microbiota of the litter floor broilers was higher, and the relative abundance levels of litter breeding bacteria (Facklamia, Globicatella, and Jeotgalicoccus) and potential pathogenic bacteria (Streptococcus and Staphylococcus) were higher (P < 0.05). Through Spearman correlation analysis, it was found that enriched microbes in the ileum of litter floor broilers were positively correlated with immune function. In summary, compared with cage broilers, litter floor broilers had more potential pathogenic bacteria and litter breeding bacteria in the ileum, which may be one of the important reasons for the stronger immune function status. IMPORTANCE In China, the three-dimensional rearing system (cage) for broilers has gradually become a trend. In production, it was found that the incidence of disease in broiler chickens raised in cage systems was significantly higher than that of ground litter. Given that broilers raised on ground litter systems may be exposed to more environmental microbes, it is important to understand whether the rearing environment affects the function and status of the host immune system by altering the gut microbiota. In this study, rearing environment-derived gut microbes associated with stronger immune function in ground litter broilers were provided, which will provide new insights into strategies to target gut microbes to promote immune function and status in broilers raised in cages.

Soya saponin fails to improve the antioxidation and immune function of laying hens with antibiotics treated.

Soya saponin (SS) helps to improve antioxidant and immune function of body, and intestinal bacteria might play an important role here. In the present study, the co-occurring network of the ileal flora was analyzed with 50 mg/kg SS supplemented to the diet, and Romboutsia was found to have evolved into a dominant flora. In addition, the co-occurring network of the flora was changed with the combined antibiotic treated, and the unidentified-cyanobacteria developed into the dominant flora, whereas the relative abundance of Romboutsia was dropped. Dietary SS failed to elevate the relative abundance of Romboutsia with antibiotics treated, at the same time, it was not helpful for the antioxidant and immune function of laying hens. While dietary SS had a little help on the egg-laying performance. Intestinal bacteria did play a key role in the biological functions of SS on laying hens. In conclusion, SS failed to improve the antioxidation and immune function of laying hens with antibiotics treated.

Effects of Dietary Astragalus Polysaccharide Supplementation on the Th17/Treg Balance and the Gut Microbiota of Broiler Chickens Challenged With Necrotic Enteritis.

This study aimed to investigate the effects of dietary astragalus polysaccharide (APS) supplementation on the immune function, gut microbiota and metabolism of broiler chickens challenged with necrotic enteritis (NE). Two hundred forty Arbor Acres broiler chicks (one day old) were randomly assigned using a 2 × 2 factorial arrangement into two groups fed different levels of dietary APS (0 or 200 ppm of diet) and two disease challenge groups (control or NE challenged). The results showed that NE infection significantly increased FCR, mortality rate, Th17/Treg (Th17 cells% in blood and ileum, Th17/Treg, IL-17 and IL-17/IL-10 in blood), NO, lysozyme activity and IL-1ß in blood, intestinal immune cell proportion and activity (Tc%, Treg% and monocyte phagocytic activity in ileum), intestinal inflammatory cytokines (TLR2, NF-κB, TNF-α and IL- 6) gene expression levels, and the number of Clostridium perfringens in cecum. NE infection significantly reduced body weight gain, thymus index, lymphocyte proliferation activity in blood and ileum, villus height and V/C in jejunum, Th cells% and Mucin2 gene expression in ileum. Dietary APS supplementation significantly increased body weight, feed intake, proportion of immune cells (T cells in blood and Tc, Treg in ileum), lymphocyte proliferation activity, V/C in jejunum, and ZO-1 gene expression in ileum. Dietary APS supplementation significantly reduced FCR and mortality rate, Th17/Treg, Th17%, intestinal pathology scores, intestinal inflammatory cytokine gene expression levels, and the number of Clostridium perfringens in cecum. In addition, broilers challenged with NE significantly increased Staphylococcus and Turicibacter and reduced α diversity of microbiota in ileum. Dietary APS supplementation significantly increased α diversity, Romboutsia, Halomonas, propionic acid, butyric acid, formononetin, taurine, cholic acid and equol and downregulated uric acid, L-arginine and serotonin in ileum. Spearman's correlation analysis revealed that Romboutsia, Turicibacter, Staphylocpccus, Halomonas, Streptococcus, Escherichia-Shigella, Prevotella, uric acid, L-arginine, jerivne, sodium cholate and cholic acid were related to inflammation and Th17/Treg balance. In conclusion, APS alleviated intestinal inflammation in broilers challenged with NE probably by regulating intestinal immune, Th17/Treg balance, as well as intestinal microbiota and metabolites.

Dietary soya saponin improves the lipid metabolism and intestinal health of laying hens.

Soya saponin (SS) is a natural active substance of leguminous plant, which could improve lipid metabolism and regulate immune function. Intestinal flora might play a key role in the biological functions of SS. The objective of this study was to measure the effects of dietary SS on immune function, lipid metabolism, and intestinal flora of laying hens with or without antibiotic treated. The experiment was designed as a factorial arrangement of 3 dietary SS treatments × 2 antibiotic treatments. Birds were fed a basal diet (CON) or a low-SS diet (50 SS) containing 50 mg/kg SS, or a high-SS diet (500 SS) containing 500 mg/kg SS. Birds were cofed with or without antibiotics. The growth experiment lasted for 10 wk. Results showed that birds fed the 50 mg/kg SS diet tended to have lower abdominal fat rate. The gene expression of liver X receptor-α (LxR-α) in liver and serum total cholesterol (TC) were dropped, and the gene expression of acyl-CoA thioesterase 8 (ACOT8) in liver were upregulated. Compared with CON group, the levels of lysozyme, IL-10, and transforming growth factor (TGF-ß) in the serum were elevated as along with gene expression of IL-10, TGF-ß, and LYZ in ileum of both 50 and 500 SS group. However, the level of secretory immunoglobulin A (sIgA) and Mucin-2 in the ileum were downregulated in the 500 SS group. Additionally, Lactobacillus and Lactobacillus gasseri were the dominant bacteria in the 50 SS group, whereas the relative abundance of Lactobacillus was dropped in the 500 SS group. With combined antibiotics treatment, the α-diversity of bacteria was reduced, and the biological effects of SS were eliminated. In conclusion, the lipid metabolism, immune function, and intestinal flora of the laying hens were improved with the dietary supplementation of 50 mg/kg SS. But dietary 500 mg/kg SS had negative effects on laying hens.

Dietary folic acid supplementation improves semen quality and spermatogenesis through altering autophagy and histone methylation in the testis of aged broiler breeder roosters.

The aging phenomenon often exerts a significant reduction in the reproduction performance of aged animals. The objective of this project was to investigate the effects of dietary Folic acid (FA) supplementation on the reproductive performance of aged broiler breeder roosters. A total of 16 aged ROSS 308 broiler breeder roosters (50-week-old) were randomly divided into two groups. The treatments were basal diet (CON), a basal diet supplemented with 10 mg/kg Folic acid (FAS) for four weeks. At the end of the experiment, semen quality, histopathological studies, serum concentrations of testosterone and relative mRNA and protein expressions of testes were evaluated. The results showed that dietary FA supplementation dramatically improved semen quality of aged roosters, manifested by increasing semen volume, sperm concentration, sperm motility, and sperm membrane functional integrity. Furthermore, seminiferous tubule epithelial height (SEH) and testis scores were increased by dietary supplementation with FA. Dietary FA also remarkably augmented the transcription level of spermatogenesis-related gene (CREM, PCK2, DDX4, and GDNF). No significant differences were observed in serum concentrations of testosterone between FAS and CON groups. We noted significant upregulation Beclin-1 and ATG5 protein expressions, and the ratio of LC3-Ⅱ/Ⅰ, as well as significant downregulation of p-mTOR protein expressions in testicular tissue of aged roosters with FA supplementation. In addition, dietary FA supplementation significantly increased the protein expression of H3K9me2 and reduced the protein expression of H3K27me2. In summary, dietary FA supplementation improved the testicular autophagy through the mTOR-signaling pathway, and altered histone methylation in the testis. Dietary supplementation with FA can ameliorate semen quality and spermatogenesis of aged roosters.

Dietary mulberry-leaf flavonoids supplementation improves liver lipid metabolism and ovarian function of aged breeder hens.

Mulberry-leaf flavonoids (MF), extracted from mulberry leaves, exert antioxidant and hypolipidemic effects. The purpose of this experimental study was to investigate the effects of dietary MF on the ovarian function and liver lipid metabolism of aged breeder hens. We used 270 (60-weeks-old) Qiling breeder hens randomly assigned in 3 treatments with supplemental dietary MF doses (0, 30, 60 mg/kg). The results showed that dietary MF significantly improved the egg-laying rate, followed by the reduced feed conversion rate (FCR) (p < 0.05). However, there is no obvious difference in hatchability and fertilised eggs hatchability among the three groups (p > 0.05). The level of T-CHO, LDL-C and AKP in serum was reduced, and the HDL-C concentrations were increased by dietary MF (p < 0.05). MF treatment also improved the antioxidant capacity and reduced the apoptotic index of the ovary (p < 0.05). Additionally, dietary MF significantly increased the serum estradiol (E2) levels (p < 0.05) and the transcription level of CYP19A1 and LHR in the ovary (p < 0.05). Dietary MF enhanced fatty acid ß-oxidation in the liver via up-regulating the mRNA expressions of PPARα and CPT-I (p < 0.05). Moreover, the HMF group significantly decreased mRNA expressions of SREBP-1c (p < 0.05) and increased mRNA expressions of ERα, VTG-Ⅱ and ApoB in the liver (p < 0.05). In conclusion, dietary MF could improve the reproduction performance of aged breeder hens through improving ovary function and hepatic lipid metabolism.