Enzymolytic soybean meal-impact on growth performance, nutrient digestibility, antioxidative capacity, and intestinal health of weaned piglets.

Enzymolytic soybean meal (ESBM) enriches free amino acids and small peptides, while mitigating anti-nutritional factors. Substituting soybean meal with ESBM enhances animal performance, though optimal piglet dietary supplementation levels vary. The present study aimed to assess the impact of ESBM on the growth performance, nutrient digestibility, antioxidative capacity and intestinal health of weaned piglets. A total of 120 piglets (initial body weight, 7.0 ± 0.4 kg) were randomly allocated into 4 dietary groups, each comprising 5 replicates with 6 piglets per replicate. The control group received the basal diet, while the experimental groups were fed diets containing 2, 4% or 8% ESBM as a replacement for soybean meal over 28 days. Compared with the control group, piglets supplemented with 4% ESBM exhibited a significant increase (p < 0.05) in average daily gain and the apparent total tract digestibility of dry matter, ether extract and gross energy (p < 0.05), alongside a notable decrease (p < 0.05) in diarrhea incidence. Fed ESBM linearly increased (p < 0.05) the villus height in the ileum of piglets. The levels of superoxide dismutase and total antioxidant capacity in serum of piglets increased (p < 0.05) in the 2 and 4% ESBM groups, while diamine oxidase content decreased (p < 0.05) in the 4 and 8% ESBM group. ESBM inclusion also upregulated (p < 0.05) the expression of superoxide dismutase 1 (SOD-1), Catalase (CAT) and claudin-1 mRNA. In terms of cecal fermentation characteristics, ESBM supplementation resulted in a increase (p < 0.05) in valerate content and a linear rise (p < 0.05) in propionate, butyrate, and total short-chain fatty acids levels, accompanied by a decrease (p < 0.05) in the concentrations of tryptamine and NH3 in cecal digesta. ESBM had no discernible effect on cecal microbial composition. In summary, substitution of soybean meal with ESBM effectively improved the growth performance of piglets by enhancing nutrient digestibility, antioxidant capacity, intestinal barrier and cecal microbial fermentation characteristics, with the optimal replacement level identified at 4%.

Curcumin alleviates cecal oxidative injury in diquat-induced broilers by regulating the Nrf2/ARE pathway and microflora.

This study evaluated the alleviative effect of curcumin (CUR) on the diquat (DQ)-induced cecal injury in broilers. A total of 320 one-day-old Cobb broilers were selected and randomly divided into 4 treatments, namely control, DQ, CUR 100, and CUR150 groups. The control and DQ groups were fed a basal diet, while the CUR 100 and CUR150 groups were fed the basal diet supplemented with 100 and 150 mg/kg CUR, respectively. Each group had 8 replicates, with 10 broilers per replicate. On day 21 of the experiment, 1 broiler was selected from each replicate and intraperitoneally injected 20 mg/kg body weight of DQ for DQ, CUR 100, and CUR 150 groups. Broilers in control group received equivalent volume of saline. Broilers were euthanized 48h postinjection for tissue sampling. The results showed that DQ injection could cause oxidative stress and inflammatory reactions in the cecum, affecting the fatty acid production and flora structure, thus leading to cecum damage. Compared with the DQ group, the activity of superoxide dismutase, the level of interleukin 10, acetic acid, and total volatile fatty, and the abundance of nuclear factor E2-related factor 2, copper and zinc superoxide dismutase and catalase mRNA in the cecal mucosa of broilers in the CUR group increased significantly (P < 0.05). However, the levels of malondialdehyd, reactive oxygen species, tumor necrosis factor-alpha, and the expression of cysteine-aspartic acid protease-3 and tumor necrosis factor-alpha decreased significantly (P < 0.05) in the CUR group. In addition, CUR treatment alleviated the damage to the cecum and restored the flora structure, and Lactobacillus and Lactobacillaceae promoted the alleviative effect of CUR on DQ. In summary, CUR could alleviate the cecal injury caused by DQ-induced oxidative damage and inflammatory reactions by regulating the Nrf2-ARE signaling pathway and intestinal flora, thus protecting the cecum.

Dietary curcumin supplementation alleviates diquat-induced oxidative stress in the liver of broilers.

This study purposed to investigate the alleviating effect of dietary curcumin supplementation on oxidative stress in the liver of broilers induced by diquat. One-day-old Cobb broilers (400) were selected and randomly divided into 5 groups, with 8 replicates and 10 broilers per replicate. The control group and the diquat group were fed the basal diet, while the curcumin supplementation groups were fed the basal diet supplemented with different amounts of curcumin (50, 100, and 150 mg/kg). On d 21 of the test, 1 broiler was randomly selected from each replicate and intraperitoneally injected with 20 mg/mL of diquat solution at a dose of 1 mL/kg BW or equivalent physiological saline (for the control group). After 48 h of feeding, the selected broilers were slaughtered for analysis. The results show that diquat treatment reduced the antioxidant capacity of the liver, caused oxidative stress, and affected its lipid metabolism. However, diet supplementation using curcumin completely or partially reversed the effect of diquat on the liver of broilers. The blood alanine aminotransferase activity, total bilirubin and total protein levels, and liver Caspase-3 mRNA abundance in broilers were lower or significantly lower in the curcumin supplementation group than in the diquat group (P < 0.05). The curcumin supplementation groups had significantly higher total antioxidant capacity activity but significantly lower malondialdehyde in the liver of broilers than the diquat group (P < 0.05). The blood triglyceride level of broilers was lower or significantly lower in the curcumin supplementation groups than in the diquat group (P < 0.05). The activities of cetyl coenzyme A carboxylase in the liver were significantly lower in the 150 mg/kg curcumin supplementation groups than in the DQ group (P < 0.05). In conclusion, dietary curcumin supplementation could ameliorate the effects of diquat-induced oxidative stress on the antioxidant capacity, tissue morphology, and lipid metabolism of the liver of broilers, thus protecting the liver. The recommended dosage for broiler diets is 100 to 150 mg/kg curcumin.

Quercetagetin alleviates zearalenone-induced liver injury in rabbits through Keap1/Nrf2/ARE signaling pathway.

Introduction: This study aimed to assess the alleviative effect of quercetagetin (QG) on zearalenone (ZEN)-induced liver injury in rabbits. Methods: Ninety 41-day-old healthy Hyla rabbits were randomly assigned into three groups, including a control (fed with basic diet), ZEN addition group (fed with basic diet + 600 µg/kg ZEN), and ZEN + QG addition group (fed with basic diet + 600 µg/kg ZEN + 100 mg/kg QG), with 30 rabbits per group. The duration of the experiment was 28 days. Results: The results revealed no significant differences in the average daily gain, average daily feed intake, the gain to feed ratio and the liver, kidney and spleen organ indexes (p > 0.05) between the rabbits across the three groups. However, the sacculus rotundus index of the rabbits in the control group was significantly higher than that in the ZEN + QG group (p < 0.05). The intake of ZEN-contaminated diet also significantly increased the activities or levels of alanine transaminase, alkaline phosphatase, total bile acid (TBA), total bilirubin, malondialdehyde, and interleukin-4 (IL-4) and enhanced the abundance of kelch-like ECH-associated protein 1 (Keap1), heat shock protein 70 (HSP70) and cysteine-aspartic acid protease-3 (Caspase-3) mRNA in the blood or liver tissue in ZEN group, compared to the control group (p < 0.05). On the contrary, the activities or levels of immunoglobulin A, complement 3, total antioxidant capacity, glutathione peroxidase (GSH-Px), superoxide dismutase, interleukin-10, and the abundance of nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) mRNA were significantly decreased (p < 0.05). Supplementing the diet with QG still maintained significantly higher levels of TBA and IL-4, and the abundance of GSH-Px, HSP70, IL-4, and Caspase-3 mRNA in the blood and liver of rabbits in the ZEN + QG group than in the control group (p < 0.05). At the same time, the other indicators were restored to levels in the control group (p > 0.05). Discussion: In conclusion, QG alleviated the ZEN-induced oxidative damage and liver injury caused by inflammatory reaction through the Keap1-Nrf2-antioxidant response element (ARE) signal pathway, which protected the liver. This study revealed the alleviative effect of QG on the hepatotoxicity of ZEN in rabbits for the first time, providing a new perspective for applying QG and developing a ZEN antidote.

Effects of dietary supplementation with quercetagetin on nutrient digestibility, intestinal morphology, immunity, and antioxidant capacity of broilers.

Quercetagetin (QG) is gaining increased attention as a potential alternative to in-feed antioxidants due to its antioxidant activity. This experiment was conducted to investigate the effects of dietary supplementation with QG on nutrient digestibility, intestinal morphology, immunity, and antioxidant capacity of broilers. Four hundred 1-day-old Ross 308 broilers were randomly assigned into 4 groups with 10 replicates in each group and 10 broilers in each replicate. The four dietary treatments included the basal diet supplemented with 0, 3.2, 4.8, or 6.4 mg/kg QG. The results showed that dietary supplementation with QG significantly promoted the broilers' apparent digestibility of phosphorus (P < 0.05), increased the villus height in jejunum and ileum, and reduced the crypt depth in jejunum and ileum, which significantly increased the ratio of villus height to crypt depth in the jejunum and ileum (P < 0.05). The dietary supplementation with QG also significantly enhanced the immunoglobulin G (IgG) and complement 4 (C4) levels in the blood (P < 0.05), the activity of total antioxidant capacity (T-AOC) in serum, jejunum mucosa, and ileum mucosa, the activity of superoxide dismutase (SOD) in the serum and liver (P < 0.05), and significantly up-regulated the kelch-like ECH-associated protein 1 (Keap1), nuclear factor E2 related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NAD(P)H: quinone oxidoreductase 1 (NQO-1), glutathione peroxidase (GSH-Px) and superoxide dismutase 1 (SOD1) mRNA expression levels in the jejunum mucosa, ileum mucosa, and liver tissues of broilers. Therefore, supplementing broilers' diets with QG can enhance the apparent digestibility of phosphorus, improve the structure and morphology of jejunum and ileum, promote immunity, and increase the activity of antioxidant enzymes and the antioxidantive capacity through the Nrf2/antioxidant response element (ARE) signaling pathway mediated by Keap1.

Effects of ammonia on hypothalamic-pituitary-ovarian axis in female rabbits.

BACKGROUND: As one of the most harmful gases in the livestock house, ammonia is recognized as an environmental stressor by Environmental Protection Agency (United States). The study aimed to explore the effect of ammonia on hypothalamic-pituitary-ovarian (HPO) axis of rabbits. A total of ninety two-month-old female IRA rabbits were randomly divided into three groups, and were kept in animal environment control rooms for four weeks at college of animal science and technology, Hebei Agricultural University (Baoding, China). The rabbits in the control group were kept under ammonia concentration of < 3 ppm. The two treatment groups were kept under ammonia concentration of 30 ppm and 50 ppm. Hypothalamus, pituitary, and ovary were collected for hematoxylin and eosin (HE) staining, immunohistochemistry, terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling (TUNEL) and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Serum was collected for enzyme-linked immunosorbent assay (ELISA). RESULTS: Histopathological examination revealed that exposed to excess ammonia damaged the morphology and structure of hypothalamus, pituitary, and ovary. TUNEL assay revealed that apoptosis rate increased in hypothalamus, pituitary, and ovary. The protein expression levels of Bcl-2associated X protein (Bax) and Caspase-9 increased, while B-cell lymphoma-2 (Bcl-2) decreased, resulting in apoptosis. Moreover, the concentration of gonadotropin-releasing hormone (GnRH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), and progesterone (PROG) reduced in plasma. The mRNA expression of FSH and LH in pituitary and follicle-stimulating hormone receptor (FSHR), E2, PROG in ovary as well as decreased, indicated hormone secretion disorder. CONCLUSIONS: The results indicated that ammonia exposure damaged hypothalamus, pituitary, and ovary, caused hormone secretion disorder and apoptosis.

Effects of ammonia on growth performance, lipid metabolism and cecal microbial community of rabbits.

This study was designed to investigate the effect of ammonia on growth performance, lipid metabolism and intestinal flora of rabbits. A total of 150 female IRA rabbits (35-days-old) were randomly divided into three groups including 0 ppm (CG), 10 ppm (LAC) and 30 ppm ammonia (HAC) groups for a period of 28 days. The average daily weight gain (ADG) of rabbits was significantly reduced in LAC (-17.11%; p < 0.001) and HAC groups (-17.46%; p < 0.001) as compared with the CG. Serum concentration of high density lipoprotein (HDL) and glucose (Glu) were increased in LAC (+80.95%; +45.99; p < 0.05) and HAC groups (+219.05%; +45.89; p < 0.001), while apolipoprotein A1 (apoA1) was decreased in LAC (-58.49%; p < 0.001) and HAC groups (-36.92%; p < 0.001). The structural integrity of cecum was damaged, and the thickness of mucosa and serosa were significantly decreased in LAC and HAC. The acetate, butyrate and propionate level of cecal chyme were reduced in HAC group (-21.67%; -19.82%; -30.81%; p < 0.05). Microbial diversity and burden of Firmicutes were significantly decreased, while that of pathogenic bacteria, such as Bacteroidetes, Clostridium and Proteobacteria were increased in ammonia treated groups. Spearman's correlation confirmed that burden of Ruminococcaceae_NK4A214_group showed significantly negative correlation with acetic acid (r = -0.67; p < 0.001) while Barnesiellaceae_unclassified showed significantly positive correlation with propionic acid (r = 0.50; p < 0.001). In conclusion, ammonia treatment was responsible for an imbalance of intestinal flora, which affected lipid metabolism and damaged intestinal barrier of rabbits, resulting in low growth performance due to lipid metabolism dysfunction.

Chicken Egg Yolk Antibody (IgY) Protects Mice Against Enterotoxigenic Escherichia coli Infection Through Improving Intestinal Health and Immune Response.

Chicken egg yolk antibody (IgY), considered as a potential substitute for antibiotics, has been used for preventing pathogens infection in food, human and animals. This study investigated effects of IgY on growth, adhesion inhibitory and morphology of enterotoxigenic Escherichia coli (ETEC) K88 in vitro, and evaluated the protective effects of IgY on intestinal health and immune response of mice infected with ETEC in vivo. Sixty pathogen-free C57BL/6J (4-6 weeks of age) mice were divided into six treatments: control (neither IgY nor ETEC infection), ETEC infection, ETEC-infected mice treated with 250 µL of high-dose (32 mg/mL), medium-dose (16 mg/mL) or low-dose (8 mg/mL) anti-ETEC IgY, or ETEC-infected mice treated with 250 µL of non-specific IgY (16 mg/mL). Anti-ETEC IgY inhibited ETEC growth, reduced adherence of ETEC to intestinal epithelial cells J2 and damaged the morphology and integrity of ETEC cell. Oral administration of anti-ETEC IgY effectively ameliorated ETEC-induced clinical signs, reduced ETEC colonization and intestinal permeability, alleviated inflammatory response through reducing the production and expression of proinflammatory cytokines, improved intestinal morphology, and inhibited excessive activation of the mucosal immune response of challenged mice. The overall protective effects of high-dose and medium-dose anti-ETEC IgY against ETEC infection were more effective. These results suggest that anti-ETEC IgY may function as a promising novel prophylactic agent against enteric pathogens infection.

Effect of gaseous hydrogen sulphide on growth performance and cecal microbial diversity of weaning pigs.

The purpose of this study was to examine the effect of gaseous hydrogen sulphide on growth performance and cecal microbial diversity in weaning pigs. A total of 24 weaning pigs (Landrace × Yorkshire × Duroc; average body weight = 8.55 ± 0.68 kg；weaning at 28 days) were selected and randomly divided into four groups (six replicates in each group). The piglets were exposed to hydrogen sulphide (0, 5, 10 and 15 mg/m3 ) during the experiment period, which lasted 28 days in four controlled environmental chambers. The results showed that exposure to hydrogen sulphide reduced the average daily gain (ADG), average daily feed intake (ADFI), and increased the diarrhoea rate of piglets. Hydrogen sulphide could increase the abundance and diversity of intestinal microbiota. The abundance of Firmicutes and Proteobacteria increased and Bacteroides decreased in the treatment groups. Five biomarkers, such as Eubacterium_1coprostanoligenes, Clostridiales, Phascolarctobacterium, Acidaminococcaceae and Ruminococcaceae_UCG_002 were selected by Lefse analysis. Our results reveal that hydrogen sulphide damaged the growth performance and destroyed the microbial bacteria balance of weaning pigs. The concentrations of hydrogen sulphide should fall below 5 mg/m3 .

Effects of zearalenone on genital organ development, serum immunoglobulin, antioxidant capacity, sex hormones and liver function of prepubertal gilts.

The study aimed to examine the effects of zearalenone on genital organ development, serum immunoglobulin, antioxidant capacity, sex hormones and liver function of prepubertal gilts. Forty-eight prepubertal gilts (Landrace × Yorkshire) were randomly divided into three treatment (T1, T2 and T3) groups and a control group (12 replicates per group, 1 gilt per replicate). Prepubertal gilts in the control group were fed with basal diet, and those in T1, T2 and T3 groups were fed with basal diets supplemented with 200 µg/kg, 800 µg/kg and 1600 µg/kg zearalenone during the experiment period, which lasted for 14 d. Feed intake was counted and vulvar area was measured. The blood samples were collected from the anterior vena cava of 6 prepubertal gilts in each group, and immunoglobulins, antioxidant indexes, inflammatory cytokines, genital hormones, and biochemical indexes were analyzed by enzyme-linked immunosorbent assay. The results showed that the average daily feed intake of prepubertal gilts in each group had no significant change (p > 0.05). On 14 d, compared with the control group, the vulva area of prepubertal gilts in each treatment group was significantly increased (p < 0.05). Compared with the control group, the serum immunoglobulin G content in the T3 group was significantly reduced (p < 0.05). The activities of total antioxidant capacity and the superoxide dismutase of serum in the T3 group were significantly reduced (p < 0.05). Compared with the control group, the serum interleukin-4 content in each test group were extremely significantly increased (p < 0.01). The serum contents of luteinizing hormone in the T2 and T3 groups and estradiol in the T3 group were significantly reduced (p < 0.05) than that of control group. Compared with the control group, the activity of aspartate aminotransferase in T3 group was significantly increased (p < 0.05). In conclusion, zearalenone has no significantly effect on the feed intake of prepubertal gilts, but it can reduce its serum immunoglobulin contents and antioxidant properties, disrupt the secretion of sex hormones, increase the vulva area, produce reproductive toxicity and cause liver damage. Therefore, in pig production, the use of antimould reagent together with products of immunity-boosting, antioxidant, anti-inflammatory and hepatoprotective may enhance protection.

Effects of dietary crude protein level and N-carbamylglutamate supplementation on nutrient digestibility and digestive enzyme activity of jejunum in growing pigs.

Three experiments were conducted to investigate the effects of dietary crude protein (CP) level and N-carbamylglutamate (NCG) supplementation on apparent total tract digestibility (ATTD) and ileal digestibility of nutrients and digestive enzyme activity of jejunum in growing pigs. In experiment 1, 10 Duroc × Landrace × Yorkshire barrows (initial BW: 48.7 kg) were allotted to a three-period switchback design with five experimental diets and two replicate pigs per diet in each period. Diets were categorized as high CP (HP, 18% CP), moderate low CP (MLP, 15% CP), very low CP (VLP, 12% CP), and MLP and VLP with 0.1% NCG supplementation. Feces and urine were collected from day 6 to day 11 after a 5-d adaptation period. The DE, ME, and ATTD of GE, OM, CP, NDF, ADF, and P decreased (P < 0.01) with a reduction of dietary CP, but no effect of dietary treatments on pig daily N retention was detected. The NCG supplementation increased (P < 0.01) DE and ATTD of ADF of the VLP diet. In experiment 2, 10 jejunal-cannulated Duroc × Landrace × Yorkshire barrows (initial BW: 44.5 kg) were fed five diets for three periods as experiment 1. Jejunal fluid was collected on days 6 and 8 after a 5-d adaptation period. The digestive enzymes activity was not affected by dietary CP level, except for α-amylase, for which there was a decrease (P < 0.01) in pigs fed VLP diets compared to HP and MLP diets. In experiment 3, 12 ileal-cannulated Duroc × Landrace × Yorkshire barrows (initial BW: 46.7 kg) were allotted to a three-period switchback design with six diets and two replicate pigs per diet in each period. The six experimental diets consisted of five experimental diets as experiment 1 and one N-free diet. Ileal digesta was collected from day 6 to day 8 after a 5-d adaptation period. Results indicated that apparent ileal digestibility (AID) of CP and P and ileal digestibility of Arg, His, Ile, Leu, Phe, and all dispensable AA, except Pro, decreased (P < 0.01) in pigs fed VLP diet compared to HP and MLP diets, but AID of GE, OM, EE, NDF, and ADF were not affected. The supplementation of NCG in the VLP diet increased (P < 0.01) the AID of CP and ileal digestibility of Arg, His, Leu, Phe, Val, Ser, and Tyr. In conclusion, reducing dietary CP level decreased nutrient digestibility, but improved the efficiency of dietary N utilization and reduced N emission. Moderate reduction of dietary CP level had a minimal effect on nutrient digestibility and digestive enzyme activity. Additionally, NCG supplementation plays a beneficial effect on nutrient digestion only if the dietary CP level is extremely lowered.

Gold nanoparticle-based colorimetric ELISA for quantification of ractopamine.

The work describes a gold nanoparticle-based colorimetric enzyme-linked immunosorbent assay (ELISA) for ractopamine. The ELISA is based on an indirect competitive approach. In the presence of ractopamine, gold(III) ions are oxidized by H2O2 to form red AuNPs. On the other hand, the AuNP in solution are purple-blue due to aggregation if the sample does not contain ractopamine. The absorption, best measured at 560 nm, increases linearly in the 2 to 512 ng·mL-1 ractopamine concentration range, and the detection limit is as low as 0.35 ng·mL-1 in urine. Ractopamine can also be detected visually, even in the presence of other ß-agonists and antibiotics. The results obtained by this method are consistent with those obtained by LC-MS/MS as demonstrated by analysis of sheep urine. The ELISA method described here is inexpensive, easy-to-use, and suitable for rapid screening of ractopamine in animal samples. Graphical abstract Schematic presentation of a colorimetric indirect competitive immunoassay for ractopamine. It is based on the use of catalase labeled IgG and the measurement of the absorption of red gold nanoparticles (AuNPs) that are generated by the reaction of gold ions with H2O2. In the absence of ractopamine, the solution becomes blue.

A sensitive and semi-quantitative method for determination of multi-drug residues in animal body fluids using multiplex dipstick immunoassay.

The objective of this research was to develop a multiplex dipstick immunoassay method for the simultaneous determination of multi-veterinary drug residues, such as ß-agonists, sulfonamides, and tetracyclines in milk, urine, and serum. The multiplex dipstick assay format was based on an indirect competitive approach: Three test lines (different antigens) and one control line (goat anti-mouse IgG) were located on the strip membrane. Labeled antibodies were freeze-dried in microwells. Samples did not require pretreatment and could be directly analyzed within 10 min. Threshold levels in different sample matrices were visually estimated at 0.3-0.45 ng mL(-1) for clenbuterol; 3-4 ng mL(-1) for sulfadiazine; and 4.5-6 ng mL(-1) for tetracycline, respectively. The linear relationship between the concentrations of veterinary drug residues and the Au nanoparticles plasmon absorbance allowed quantitative determination of these veterinary drug residues. The recoveries of clenbuterol, sulfadiazine and tetracycline in spiked samples ranged from 78.4% to 112.6%, and the relative standard deviations were below 11.2%. Analysis of animal samples suggested that the proposed multiplex dipstick assay method was consistent with the LC-MS/MS method. The percentage of false results was less than or equal to 5%. Thus, the proposed multiplex dipstick assay is inexpensive, easy-to-use, and suitable for the purposes of rapid and comprehensive screening of 3 families of ß-agonists, sulfonamides and tetracyclines including 26 drugs in animal body fluids.