Distribution and health risk of chromium in wheat grains at the national scale in China.

Chromium (Cr) pollution may threaten food safety in China. In this study, the concentration, pollution level, distribution, and non-cancer risk of Cr in wheat grains grown in 186 areas across 28 provinces in China were investigated. Results indicated that mean concentration of Cr was 0.28 ± 2.5 mg/kg, dry mass (dm). Of the samples, 7.5 % were found to be polluted with Cr. The mean concentrations were in the following order: Northwest > Northeast > South > East > North > Southwest > Central China. Based on deterministic models, mean hazard quotient (HQ) values for adult males, adult females, and children were 0.11 ± 3.4, 0.11 ± 3.4, and 0.13 ± 3.5, respectively with < 6 % of HQ values ≥ 1. Eleven sites in northern China were identified as hotspots, whereas Gansu Province and Northwestern China were labeled as priority provinces and regions for risk control. The mean HQ values estimated by probabilistic risk assessment were two times greater than those estimated using deterministic models. The risk probabilities for adult males, adult females, and children were 4.81 %, 3.78 %, and 6.55 %, respectively. This study provides valuable information on Cr pollution in wheat grains and its risks at a national scale in China.

Predicting metabolizable energy of soybean meal and rapeseed meal from chemical composition in broilers of different ages.

This study determined metabolizable energy (ME) and developed ME prediction equations for broilers based on chemical composition of soybean meal (SBM) and rapeseed meal (RSM) using a 2 × 10 factorial arrangement of age (11 to 14 or 25 to 28 d of age) and 10 sources of each ingredient. Each treatment contained 6 replicates of 8 broilers. The ME values were determined by total collection of feces and urine. Principal components analysis (PCA) of the chemical composition clearly revealed distinct differences in SBM and RSM based on a principal components (PC) score plot. The nitrogen-corrected apparent metabolizable energy (AMEn) of SBM was higher in broilers from 25 to 28 than 11 to 14 d of age (P = 0.013). Interactions between broiler age and ingredient source affected apparent metabolizable energy (AME) of SBM and ME of RSM (P < 0.05). The ME of SBM in 11 to 14 and 25 to 28-day-old broilers were estimated by crude protein (CP) content (R2≥ 0.782; SEP ≤ 83 kcal/kg DM; P < 0.001). The AME and AMEn of RSM in 11 to 14-day-old broilers were estimated by ether extract (EE), ash and acid detergent fiber (ADF) (R2 = 0.897, SEP = 106 kcal/kg DM; P = 0.002), and by EE and ash (R2 = 0.885, SEP = 98 kcal/kg DM; P = 0.001), respectively. The AME and AMEn of RSM in 25 to 28-day-old broilers were estimated by ash and ADF (R2 = 0.925, SEP = 104 kcal/kg DM; P < 0.001) and by ash and neutral detergent fiber (NDF) (R2 = 0.921, SEP = 91 kcal/kg DM; P < 0.001), respectively. These results indicate that ME of these 2 plant protein ingredients are affected interactively by chemical composition and age of broilers. This study developed robust, age-specific prediction equations of ME for broilers based on chemical composition for SBM and RSM. Overall, ME values can be predicted from CP content for SBM, or EE, ash, ADF, and NDF for RSM.

Zinc alleviates thermal stress-induced damage to the integrity and barrier function of cultured chicken embryonic primary jejunal epithelial cells via the MAPK and PI3K/AKT/mTOR signaling pathways.

Zinc (Zn) could alleviate the adverse effect of high temperature (HT) on intestinal integrity and barrier function of broilers, but the underlying mechanisms remain unclear. We aimed to investigate the possible protective mechanisms of Zn on primary cultured broiler jejunal epithelial cells exposed to thermal stress (TS). In Exp.1, jejunal epithelial cells were exposed to 40℃ (normal temperature, NT) and 44℃ (HT) for 1, 2, 4, 6, or 8 h. Cells incubated for 8 h had the lowest transepithelial resistance (TEER) and the highest phenol red permeability under HT. In Exp.2, the cells were preincubated with different Zn sources (Zn sulfate as iZn and Zn proteinate with the moderate chelation strength as oZn) and Zn supplemental levels (50 and 100 µmol/L) under NT for 24 h, and then continuously incubated under HT for another 8 h. TS increased phenol red permeability, lactate dehydrogenase (LDH) activity and p-PKC/PKC level, and decreased TEER, cell proliferation, mRNA levels of claudin-1, occludin, zona occludens-1 (ZO-1), PI3K, AKT and mTOR, protein levels of claudin-1, ZO-1 and junctional adhesion molecule-A (JAM-A), and the levels of p-ERK/ERK, p-PI3K/PI3K and p-AKT/AKT. Under HT, oZn was more effective than iZn in increasing TEER, occludin, ZO-1, PI3K, and AKT mRNA levels, ZO-1 protein level, and p-AKT/AKT level; supplementation with 50 µmol Zn/L was more effective than 100 µmol Zn/L in increasing cell proliferation, JAM-A, PI3K, AKT, and PKC mRNA levels, JAM-A protein level, and the levels of p-ERK/ERK and p-PI3K/PI3K; furthermore, supplementation with 50 µmol Zn/L as oZn had the lowest LDH activity, and the highest ERK, JNK-1, and mTOR mRNA levels. Therefore, supplemental Zn, especially 50 µmol Zn/L as oZn, could alleviate the TS-induced integrity and barrier function damage of broiler jejunal epithelial cells possibly by promoting cell proliferation and tight junction protein expression via the MAPK and PI3K/AKT/mTOR signaling pathways.

Dietary trace mineral pattern influences gut microbiota and intestinal health of broilers.

Dietary trace minerals can impact gut flora, which can further affect intestinal health. However, the dietary balance pattern of trace minerals for the intestinal health of broilers needs to be explored. The present study was conducted to investigate the effect of the dietary pattern of Cu, Fe, Mn, Zn, and Se on the intestinal morphology, microbiota, short-chain fatty acid concentrations, antioxidant status, and the expression of tight junction proteins in broilers. A total of 240 1-d-old Arbor Acres male broilers were randomly assigned to one of five treatments with six replicate cages of eight birds per cage for each treatment. The birds were fed the corn-soybean meal basal diet supplemented with five combination patterns of trace minerals for 42 d. The dietary treatments were as follows: the inorganic sources were added to the diet based on the recommendations of the current National Research Council (NRC, T1) and Ministry of Agriculture of P.R. China (MAP) (T2) for broiler chicks, respectively; the inorganic sources were added to the diet at the levels based on our previous results of inorganic trace mineral requirements for broilers (T3); the organic sources were added to the diet at the levels considering the bioavailabilities of organic trace minerals for broilers described in our previous studies (T4); and the organic sources were added to the diet based on the recommendations of the current MAP for broiler chicks (T5). The results showed that broilers from T1 had lower (P < 0.05) crypt depth (CD), and a higher (P < 0.05) villus height: CD in duodenum on day 21 and lower CD (P < 0.05) in jejunum on day 42 than those from T3 and T4. Broilers from T1, T3, and T5 had a higher (P < 0.05) Shannon index in cecum on day 21 than those from T4. Broilers from T1 had a higher (P < 0.05) abundance of Lactobacillus in ileum on day 21 than those from T2 and T3. Broilers from T1, T2, and T5 had a higher (P < 0.05) valeric acid concentrations in cecum on day 42 than those from T3 and T4. In addition, Birds from T2 had higher (P < 0.05) Claudin-1 mRNA levels in jejunum on day 42 than those from T3 and T4. And birds from T3, T4, and T5 had a higher (P < 0.05) Occludin protein expression levels in duodenum on day 42 than those from T2. These results indicate that dietary pattern of Cu, Fe, Mn, Zn, and Se influenced gut flora and intestinal health of broilers, and the appropriate pattern of Cu, Fe, Mn, Zn, and Se in the diet for intestinal health of broilers would be Cu 12 mg, Fe 229 mg, Mn 81 mg, Zn 78 mg, and Se 0.24 mg/kg (1 to 21 d of age), and Cu 11 mg, Fe 193 mg, Mn 80 mg, Zn 73 mg, and Se 0.22 mg/kg (22 to 42 d of age), when the trace minerals as inorganic sources were added to diets according to the recommendations of the current NRC.

Information is still scarce regarding the effect of dietary trace mineral patterns on the intestinal health of broilers. The results indicated that dietary trace mineral pattern influenced intestinal health of broilers, and the appropriate pattern of trace minerals in the diet for intestinal health of broilers would be Cu 12 mg, Fe 229 mg, Mn 81 mg, Zn 78 mg, and Se 0.24 mg/kg (1 to 21 d of age), and Cu 11 mg, Fe 193 mg, Mn 80 mg, Zn 73 mg, and Se 0.22 mg/kg (22 to 42 d of age), when the trace minerals as inorganic sources were added to diets according to the recommendations of the current National Research Council. Our results provided scientific experimental bases for improving intestinal health of broilers by nutritional strategy.

Dietary calcium and nonphosphate phosphorus interaction influences tibiotarsus development and related gene expression of broilers from 1 to 21 days of age.

The dietary needs of calcium (Ca) and phosphorus (P) are interdependent, thus accurate evaluation of Ca and P requirements of broilers to support skeleton health and optimal growth is critical. The present study was carried out to investigate the effects of dietary Ca and nonphytate P (NPP) levels and their interactions on growth performance, tibiotarsus characteristics, tibiotarsus metabolism-related enzyme and proteins, and their gene expression of broilers, so as to provide a rational recommendation for Ca and NPP levels in diet. A total of 540 one-day-old Arbor Acres male broilers were randomly allotted to 1 of 15 treatments with 6 replicate cages of 6 birds per cage for each treatment in a completely randomized design involving a 5 × 3 factorial arrangement of treatments (5 levels of Ca × 3 levels of NPP). The birds were fed the corn-soybean meal diet containing 0.60%, 0.70%, 0.80%, 0.90%, or 1.00% Ca and 0.35%, 0.40%, or 0.45% NPP for 21 d. Dietary Ca level affected (P < 0.03) the bone mineral density, bone mineral content (BMC), breaking strength, ash percentage and ash Ca contents in tibia, which showed linear (P < 0.006) responses to dietary Ca levels. Dietary NPP level affected (P < 0.05) tibia BMC, ash percentage, and FGF23 mRNA level. Broilers that received 0.40% and 0.45% NPP had higher (P < 0.04) tibia BMC and ash percentage than those that received 0.35% NPP, but no differences (P > 0.05) were found between 0.40% and 0.45% NPP. Broilers that received 0.40% NPP had higher (P = 0.02) tibia FGF23 mRNA level than those that received 0.35% NPP, but no differences (P > 0.05) were detected between 0.40% and 0.45% NPP or 0.45% and 0.35% NPP. The interactions between dietary Ca and NPP affected (P < 0.05) ADG, ALP activity, bone gal protein, FGF23 contents, and the mRNA expression levels ALP and bone gal protein in tibia of broilers. Results from the present study indicate that dietary Ca and NPP interaction influences growth, tibiotarsus development, and related gene expression of broiler chickens. Considering all the criteria, the dietary levels of 0.90% Ca and 0.45% NPP would be optimal for both growth and tibiotarsus development of broilers fed a conventional corn-soybean meal diet from 1 to 21 d of age.

Manganese alleviates heat stress of primary cultured chick embryonic myocardial cells via enhancing manganese superoxide dismutase expression and attenuating heat shock response.

Manganese (Mn) is an essential trace element that has been shown to attenuate the adverse effects of heat stress in the heart of broiler breeders and embryos. However, the underlying molecular mechanisms involving this process remain unclear. Therefore, two experiments were conducted to investigate the possible protective mechanisms of Mn on primary cultured chick embryonic myocardial cells exposed to heat challenge. In experiment 1, the myocardial cells were exposed to 40 °C (normal temperature, NT) and 44 °C (high temperature, HT) for 1, 2, 4, 6 or 8 h. In experiment 2, the myocardial cells were preincubated with no Mn supplementation (CON), 1 mmol/L of Mn as the inorganic MnCl2 (iMn) or organic Mn proteinate (oMn) under NT for 48 h, and then continuously incubated under NT or HT for another 2 or 4 h. The results from experiment 1 showed that the myocardial cells incubated for 2 or 4 h had the highest (P < 0.0001) heat-shock protein 70 (HSP70) or HSP90 mRNA levels than those incubated for other incubation times under HT. In experiment 2, HT increased (P < 0.05) the heat-shock factor 1 (HSF1) and HSF2 mRNA levels as well as Mn superoxide dismutase (MnSOD) activity of myocardial cells compared with NT. Furthermore, supplemental iMn and oMn increased (P < 0.02) HSF2 mRNA level and MnSOD activity of myocardial cells compared with the CON. Under HT, the HSP70 and HSP90 mRNA levels were lower (P < 0.03) in iMn group than in the CON group, in oMn group than in iMn group; and the MnSOD mRNA and protein levels were higher (P < 0.05) in oMn group than in the CON and iMn groups. These results from the present study indicate that supplemental Mn, especially oMn, could enhance the MnSOD expression and attenuate heat shock response to protect against heat challenge in primary cultured chick embryonic myocardial cells.

The organic zinc with moderate chelation strength enhances the expression of related transporters in the jejunum and ileum of broilers.

Our previous study demonstrated that the zinc (Zn) proteinate with moderate chelation strength (Zn-Prot M) enhanced the Zn absorption in the small intestine partially via increasing the expression of some Zn and amino acid transporters in the duodenum of broilers. However, it remains unknown whether the Zn-Prot M could also regulate the expression of related transporters in the jejunum and ileum of broilers in the above enhancement of Zn absorption. The present study was conducted to investigate the effect of the Zn-Prot M on the expression of related transporters in the jejunum and ileum of broilers compared to the Zn sulfate (ZnS). Zinc-deficient broilers (13-d-old) were fed with the Zn-unsupplemented basal diets (control) or the basal diets supplemented with 60 mg Zn/kg as ZnS or Zn-Prot M for 26 d. The results showed that in the jejunum, compared to the control, supplementation of the organic or inorganic Zn increased (P < 0.05) mRNA and protein expression of b0,+-type amino acid transporter (rBAT), Zn transporter 10 (ZnT10), and peptide-transporter 1 (PepT1) mRNA expression and Zn transporter 7 (ZnT7) protein expression on d 28, while y+L-type amino transporter 2 (y+LAT2) mRNA and protein expression, and protein expression of ZnT7 and ZnT10 on 28 d and zrt-irt-like protein 3 (ZIP3) and zrt-irt-like protein 5 (ZIP5) on d 39 were higher (P < 0.05) for Zn-Prot M than for ZnS. In the ileum, Zn addition regardless of Zn source up-regulated (P < 0.05) mRNA expression of Zn transporter 9 (ZnT9) and ZIP3, ZIP5, and y+LAT2 protein expression on d 28, and PepT1 mRNA and protein expression, ZIP3 and y+LAT2 mRNA expression and ZnT10 protein expression on d 39. Furthermore, Zn transporter 4 (ZnT4) and ZnT9 mRNA expression and Zn transporter 1 (ZnT1) protein expression on d 28, and y+LAT2 mRNA expression and ZnT10 and PepT1 protein expression on d 39 were higher (P < 0.05) for Zn-Prot M than for ZnS. It was concluded that the Zn-Prot M enhanced the expression of the ZnT1, ZnT4, ZnT9, ZnT10, ZIP3, ZIP5, y+LAT2, and PepT1 in the jejunum or ileum of broilers compared to the ZnS.

Metabolic utilization of intravenously injected iron from different iron sources in target tissues of broiler chickens.

No information is available regarding the utilization of iron (Fe) from different Fe sources at a target tissue level. To detect differences in Fe metabolic utilization among Fe sources, the effect of intravenously injected Fe on growth performance, hematological indices, tissue Fe concentrations and Fe-containing enzyme activities and gene expressions of Fe-containing enzymes or protein in broilers was investigated. On d 22 post-hatching, a total of 432 male chickens were randomly allotted to 1 of 9 treatments in a completely randomized design. Chickens were injected with either a 0.9% (wt/vol) NaCl solution (control) or a 0.9% NaCl solution supplemented with Fe sulphate or 1 of 3 organic Fe sources. The 3 organic Fe sources were Fe chelates with weak (Fe-MetW), moderate (Fe-ProtM) or extremely strong (Fe-ProtES) chelation strength. The 2 Fe dosages were calculated according to the Fe absorbabilities of 10% and 20% every 2 d for a duration of 20 d. Iron injection did not affect (P > 0.05) ADFI, ADG or FCR during either 1 to 10 d or 11 to 20 d after injections. Hematocrit and Fe concentrations in the liver and kidney on d 10 after Fe injections, and Fe concentrations in the liver or pancreas and ferritin heavy-chain (FTH1) protein expression level in the liver or spleen on d 20 after Fe injections increased (P ≤ 0.05) as injected Fe dosages increased. When the injected Fe level was high at 20% Fe absorbability, the chickens injected with Fe-ProtES had lower (P < 0.001) liver or kidney Fe concentrations and spleen FTH1 protein levels than those injected with Fe-MetW or Fe-ProtM on d 20 after injections. And they had lower (P < 0.05) liver cytochrome C oxidase mRNA levels on d 20 after injections than those injected with Fe-MetW or Fe sulphate. The results from this study indicate that intravenously injected Fe from Fe-ProtES was the least utilizable and functioned in the sensitive target tissue less effectively than Fe from Fe sulfate, Fe-MetW or Fe-ProtM.

Effect of in ovo manganese injection on the embryonic development, antioxidation, hatchability, and performances of offspring broilers under normal and high temperatures.

Two experiments were carried out to study the effect of in ovo manganese (Mn) injection on the embryonic development, antioxidation, hatchability, and performances of offspring broilers under normal temperature (NT) and high temperature (HT). Experiment 1 was conducted to investigate the effect of in ovo Mn injection on the embryonic hatchability of Arbor Acres broiler breeders. On D 9 of incubation, a total of 684 fertilized eggs were randomly allocated to 6 treatments: the non-injected positive control (niPC) and treatments injected with 0 (the negative control, iNC), 6.25, 12.5, 25.0, or 50.0 µg Mn/egg as Mn sulfate. Experiment 2 was conducted to investigate the effect of in ovo Mn injection on the embryonic development, antioxidation and performance of offspring broilers under NT and HT. A total of 792 fertilized eggs were randomly allocated to 6 treatments in a 1 (niPC) + 1 (iNC) + 2 (injected Mn sources: Mn sulfate and Mn proteinate) × 2 (injected Mn levels: 12.5 and 25.0 µg/egg) factorial arrangement during the embryonic stage and D1 to 28 at NT. Then, 288 birds were allotted to 12 treatments in a 6 (the above embryonic treatments) × 2 (environmental temperatures: NT-22℃ vs HT-34℃) factorial arrangement from D 29 to 42. The results showed that Mn injection affected (P < 0.03) the hatchability and the maximum level of in ovo injected Mn was 25.0 µg Mn/egg. The Mn injection upregulated (P < 0.05) Mn-containing superoxide dismutase mRNA expression in the embryonic heart compared to the iNC. Hyperthermia decreased (P < 0.05) ADG and ADFI, breast muscle percentage, plsma alkaline phosphatase activity, and red color values of breast and thigh muscles, but increased (P < 0.05) F/G, plasma aspartate aminotransferase and lactic dehydrogenase activities, total cholesterol, uric acid and triiodothyronine contents, abdominal fat, light values of breast and thigh muscles of offspring broilers. The results suggest that in ovo Mn injection can enhance antioxidant ability in the chick embryonic heart.

Research Note: Metabolic utilization of iron from different iron sources in primary cultured hepatocytes of broiler embryos.

The present study was carried out to evaluate the effects of iron (Fe) sources and levels on the Fe concentration and expressions of iron-containing enzymes or protein in primary cultured hepatocytes of broiler embryos. The hepatocytes were incubated with 0, 0.25 and 0.50 mmol/L added Fe from either Fe sulfate, or 1 of 3 organic Fe chelates with weak (Fe-Met W), moderate (Fe-Pro M), or extremely strong (Fe-Pro ES) chelation strengths for 24 h. The results showed that all supplemental Fe treatments had higher (P < 0.05) Fe concentration, succinate dehydrogenase (SDH), CAT and ferritin heavy chain 1 (FTH1) mRNA levels than those in the control group. The hepatocytes incubated with Fe-Prot ES had lower (P < 0.009) Fe concentration than those incubated with Fe sulfate, Fe-Met W or Fe-Prot M. The SDH mRNA level was lower (P < 0.05) in Fe sulfate and Fe-Prot ES groups than in Fe-Prot M group. In conclusion, the Fe from Fe-Prot ES was less utilizable than Fe from Fe sulfate, Fe-Met W or Fe-Pro M in primary cultured hepatocytes of broiler embryos.

The Effects of Inorganic Phosphorus Levels on Phosphorus Utilization, Local Bone-Derived Regulators, and BMP/MAPK Pathway in Primary Cultured Osteoblasts of Broiler Chicks.

Understanding the underlying mechanisms that regulate the bone phosphorus (P) utilization would be helpful for developing feasible strategies to improve utilization efficiency of P in poultry. We aimed to investigate the effects of inorganic P levels on P utilization, local bone-derived regulators and bone morphogenetic protein/mitogen-activated protein kinase (BMP/MAPK) pathway in primary cultured osteoblasts of broiler chicks in order to address whether local bone-derived regulators or BMP/MAPK pathway was involved in regulating the bone P utilization of broilers using an in vitro model. The primary cultured tibial osteoblasts of broiler chicks were randomly divided into one of five treatments with six replicates for each treatment. Then, cells were respectively incubated with 0.0, 0.5, 1.0, 1.5, or 2.0 mmol/L of added P as NaH2PO4 for 24 days. The results showed that as added P levels increased, tibial osteoblastic P retention rate, number and area of mineralized nodules, the mRNA expressions of endopeptidases on the X chromosome (PHEX), dentin matrix protein 1 (DMP1), bone morphogenetic protein 2 (BMP2), and the mRNA and protein expressions of matrix extracellular phosphoglycoprotein (MEPE) increased linearly (p < 0.001) or quadratically (p < 0.04), while extracellular signal-regulated kinase 1 (ERK1) mRNA expression and c-Jun N-terminal kinase 1 (JNK1) phosphorylated level decreased linearly (p < 0.02) or quadratically (p < 0.01). Correlation analyses showed that tibial osteoblastic P retention rate was positively correlated (r = 0.452-0.564, p < 0.03) with MEPE and BMP2 mRNA expressions. Furthermore, both number and area of mineralized nodules were positively correlated (r = 0.414-0.612, p < 0.03) with PHEX, DMP1, MEPE, and BMP2 mRNA expressions but negatively correlated (r = -0.566 to -0.414, p < 0.04) with the ERK1 mRNA expression and JNK1 phosphorylated level. These results suggested that P utilization in primary cultured tibial osteoblasts of broiler chicks might be partly regulated by PHEX, DMP1, MEPE, BMP2, ERK1, and JNK1.

Evaluation of optimal dietary calcium level by bone characteristics and calcium metabolism-related gene expression of broilers from 22 to 42 d of age.

The current dietary Ca recommendation of broilers is primarily based on the previous studies carried out more than 30 yr ago. However, the modern commercial broilers are quite different from those more than 30 yr ago. The present experiment was conducted to evaluate an optimal dietary Ca level by bone characteristics and Ca metabolism-related gene expression of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 252 22-d-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicate cages of 6 birds per cage for each treatment. Broilers were fed the corn-soybean meal diets containing 0.50%, 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, or 1.10% Ca for 21 d, and each diet contained 0.31% non-phytate P. The results showed that the mineral contents in tibia and middle toe bone, mineral density in tibia and middle toe bone, middle toe ash percentage, middle toe ash Ca percentage, and tibia alkaline phosphatase mRNA expression level of broilers were influenced (P < 0.04) by dietary Ca level and increased quadratically (P < 0.05) as dietary Ca level increased. The estimates of optimal dietary Ca levels were 0.55%, 0.60%, 0.70%, 0.72%, 0.63%, 0.66%, and 0.70%, respectively, based on the best fitted broken-line, quadratic, or asymptotic models (P < 0.02) of the above sensitive indices. These results indicate that the optimal dietary Ca level would be 0.72% to support all of the Ca metabolism and bone development of broilers fed the corn-soybean meal diet from 22 to 42 d of age.

The present experiment was conducted to evaluate an optimal dietary Ca level by bone characteristics and Ca metabolism-related gene expression of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 252 22-d-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicate cages of 6 birds per cage for each treatment. Broilers were fed the corn-soybean meal diets containing 0.50%, 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, or 1.10% Ca for 21 d, and each diet contained 0.31% non-phytate P. The results showed that the tibia and middle toe bone mineral contents, tibia and middle toe bone mineral density, middle toe ash percentage, middle toe ash Ca percentage, and tibia alkaline phosphatase mRNA expression level were sensitive criteria to estimate the optimal dietary Ca levels of broilers. The estimates of optimal dietary Ca levels were 0.55% to 0.72% based on the above sensitive criteria. These results indicate that the optimal dietary Ca level would be 0.72% to support all of the Ca metabolism and bone development of broilers fed the corn-soybean meal diet from 22 to 42 d of age.

Dietary calcium requirements of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age.

BACKGROUND: The current calcium (Ca) recommendation for broilers is primarily based on studies conducted more than 30 years ago with birds of markedly different productive potentials from those which exist today. And the response indicators in these studies are mainly growth performance and bone ash percentage. Therefore, the present study was carried out to investigate the effect of dietary Ca level on growth performance, serum parameters, bone characteristics and Ca metabolism-related gene expressions, so as to estimate dietary Ca requirements of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age. METHODS: A total of 420 1-day-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicates (10 birds per cage) and fed the corn-soybean meal diets containing 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, 1.10% or 1.20% Ca for 21 days. Each diet contained the constant non-phytate phosphorus content of about 0.39%. RESULTS: The average daily gain decreased linearly (P < 0.001) as dietary Ca level increased. The serum and tibia alkaline phosphatase (ALP) activities, tibia bone mineral density (BMD), middle toe BMD, tibia ash percentage, tibia breaking strength, and tibia ALP protein expression level were affected (P < 0.05) by dietary Ca level, and showed significant quadratic responses (P < 0.02) to dietary Ca levels. The estimates of dietary Ca requirements were 0.80 to 1.00% based on the best fitted broken-line or quadratic models (P < 0.03) of the above serum and bone parameters, respectively. CONCLUSIONS: The results from the present study indicate that the Ca requirements would be about 0.60% to obtain the best growth rate, and 1.00% to meet all of the Ca metabolisms and bone development of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age.

Determination of dietary copper requirement by the monoamine oxidase activity in kidney of broilers from 1 to 21 days of age.

The current dietary copper (Cu) requirement (8 mg/kg) of broilers is mainly based on growth, hemoglobin concentration, or hematocrit, which might not be the most sensitive indices to evaluate dietary Cu requirements of broilers. The present study was carried out to estimate dietary Cu requirements of broilers fed a conventional corn-soybean meal diet from 1 to 21 d of age using biochemical or molecular biomarkers. A total of 384 1-d-old Arbor Acres male broilers were randomly allocated to 1 of 6 treatments with 8 replicates and fed a Cu-unsupplemented corn-soybean meal basal diet containing 5.17 mg Cu/kg by analysis and the basal diet supplemented with 3, 6, 9, 12 or 15 mg Cu/kg as CuSO4⋅5H2O for 21 d. Regression analysis was performed to estimate the optimal dietary Cu level using the broken-line model. Dietary supplemental Cu level affected (P < 0.05) Cu contents in serum and liver and kidney monoamine oxidase (MAO) activity, but had no effects (P > 0.05) on the growth performance, Cu contents in heart, kidney, pancreas and spleen, Cu- and zinc-containing superoxide dismutase (CuZnSOD) activity and ceruloplasmin content in serum, CuZnSOD and cytochrome c oxidase (COX) activities and ceruloplasmin, CuZnSOD, MAO A, MAO B, COX 4 I 1 and COX 1 mRNA and protein expressions in the above tissues of broilers. As dietary supplemental Cu levels increased, Cu contents in serum and liver increased linearly (P < 0.05), but kidney MAO activity decreased linearly and quadratically (P < 0.05). The estimated dietary Cu requirement based on the fitted broken-line model (P = 0.035) of kidney MAO activity was 11.30 mg/kg. In conclusion, kidney MAO activity is a new and sensitive criterion to evaluate the dietary Cu requirement of broilers, and the dietary Cu requirement was 11.30 mg/kg for broilers fed the conventional corn-soybean meal diet from 1 to 21 d of age, which is higher than the current National Research Council (NRC) Cu requirement (8 mg/kg) of broilers.

Determination of optimal dietary selenium levels by full expression of selenoproteins in various tissues of broilers from 22 to 42 d of age.

The current NRC dietary selenium (Se) requirement (0.15 mg/kg) of broilers from 22 to 42 d of age is primarily based on a previous study reported in 1986, which might not be applicable to modern classes of rapidly growing broilers. The present experiment was conducted to determine the optimal dietary Se level for meeting metabolic and functional Se requirements of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 336 Arbor Acres male broilers at 22 d old were randomly assigned to 1 of 6 treatments with 7 replicates and fed a basal corn-soybean meal diet (control, containing 0.014 mg Se/kg) and the basal diet supplemented with 0.10, 0.20, 0.30, 0.40, or 0.50 mg Se/kg from Na2SeO3 for 21 d. The results showed that the Se concentrations in plasma, liver, kidney, pancreas, breast and thigh muscles, the activity of glutathione peroxidase (GPX) in plasma, liver and kidney, the mRNA expression levels of Gpx4, selenoprotein (Seleno) h and Selenou in liver, S elenop and Selenoh in kidney, and the protein expression levels of GPX4 in the liver and kidney of broilers were affected (P < 0.05) by supplemental Se level, and increased quadratically (P < 0.05) with the increase of supplemental Se level. The estimates of optimal dietary Se levels were 0.10 to 0.49 mg/kg based on the fitted broken-line or asymptotic models (P < 0.0001) of the above Se concentration indices, and 0.08 to 0.37 mg/kg based on the fitted broken-line, quadratic or asymptotic models (P < 0.007) of the above selenoprotein expression indices. These results indicate that the optimal dietary Se levels would be 0.49 mg/kg to support the maximum Se concentrations and 0.37 mg/kg to support the full expression of selenoproteins in plasma and various tissues of broilers fed a corn-soybean meal diet from 22 to 42 d of age.

Novel Hybrid Peptide Cathelicidin 2 (1-13)-Thymopentin (TP5) and Its Derived Peptides with Effective Antibacterial, Antibiofilm, and Anti-Adhesion Activities.

The increasing numbers of infections caused by multidrug-resistant (MDR) pathogens highlight the urgent need for new alternatives to conventional antibiotics. Antimicrobial peptides have the potential to be promising alternatives to antibiotics because of their effective bactericidal activity and highly selective toxicity. The present study was conducted to investigate the antibacterial, antibiofilm, and anti-adhesion activities of different CTP peptides (CTP: the original hybrid peptide cathelicidin 2 (1-13)-thymopentin (TP5); CTP-NH2: C-terminal amidated derivative of cathelicidin 2 (1-13)-TP5; CTPQ: glutamine added at the C-terminus of cathelicidin 2 (1-13)-TP5) by determining the minimal inhibitory concentrations (MICs), minimal bactericidal concentrations (MBCs), propidium iodide uptake, and analysis by scanning electron microscopy, transmission electron microscopy, and confocal laser scanning microscopy). The results showed that CTPs had broad-spectrum antibacterial activity against different gram-positive and gram-negative bacteria, with MICs against the tested strains varying from 2 to 64 µg/mL. CTPs at the MBC (2 × MIC 64 µg/mL) showed strong bactericidal effects on a standard methicillin-resistant Staphylococcus aureus strain ATCC 43300 after co-incubation for 6 h through disruption of the bacterial membrane. In addition, CTPs at 2 × MIC also displayed effective inhibition activity of several S. aureus strains with a 40-90% decrease in biofilm formation by killing the bacteria embedded in the biofilms. CTPs had low cytotoxicity on the intestinal porcine epithelial cell line (IPEC-J2) and could significantly decrease the rate of adhesion of S. aureus ATCC 43300 on IPEC-J2 cells. The current study proved that CTPs have effective antibacterial, antibiofilm, and anti-adhesion activities. Overall, this study contributes to our understanding of the possible antibacterial and antibiofilm mechanisms of CTPs, which might be an effective anti-MDR drug candidate.

Determination of optimal dietary selenium levels by full expression of selenoproteins in various tissues of broilers from 1 to 21 d of age.

The current NRC dietary selenium (Se) requirement (0.15 mg/kg) of broilers is primarily based on growth performance data reported in 1986. Our study aimed to determine optimal dietary Se levels of broilers fed a practical corn-soybean meal diet for the full expression of selenoproteins in various tissues. A total of 384 one-d-old male broilers (n = 8 replicates/diet) were fed a basal corn-soybean meal diet or the basal diet supplemented with 0.1, 0.2, 0.3, 0.4 or 0.5 mg Se/kg in the form of Na2SeO3 for 21 d. Regression analysis was conducted to evaluate the optimal dietary Se levels using broken-line, quadratic or asymptotic models. The activity of glutathione peroxidase (GPX) in the plasma, liver, kidney and pancreas, iodothyronine deiodinase (DIO) in the plasma, liver and pancreas, and thioredoxin reductase (Txnrd) in the liver and pancreas, the mRNA levels of Gpx1, Gpx4, Dio1, selenoprotein (Seleno) h, Selenop and Selenou in the liver, Gpx4, Dio1, Txnrd1, Txnrd2, Selenoh, Selenop and Selenou in the kidney, and Gpx1, Gpx4, Selenoh and Selenou in the pancreas, and the protein levels of GPX4 in the liver and kidney of broilers were influenced (P < 0.05) by added Se levels, and increased quadratically (P < 0.05) with the increase of added Se levels. The estimates of optimal dietary Se levels were 0.07 to 0.36 mg/kg based on the fitted broken-line, quadratic or asymptotic models (P < 0.001) of the aforementioned selenoprotein expression in the plasma, liver and kidney, and 0.09 to 0.46 mg/kg based on the fitted broken-line models (P < 0.001) of the aforementioned selenoprotein expression in the pancreas. The results indicate that the optimal dietary Se levels would be 0.36 mg/kg to support the full expression of selenoproteins in the plasma, liver and kidney, and 0.46 mg/kg to support the full expression of selenoproteins in the pancreas of broilers fed a practical corn-soybean meal diet from 1 to 21 d of age.

Regulation of bone phosphorus retention and bone development possibly by related hormones and local bone-derived regulators in broiler chicks.

BACKGROUND: Phosphorus is essential for bone mineralization in broilers, however, the underlying mechanisms remain unclear. We aimed to investigate whether bone phosphorus retention and bone development might be regulated by related hormones and local bone-derived regulators in broilers. METHODS: Broilers were fed diets containing different levels of non-phytate phosphorus (NPP) 0.15%, 0.25%, 0.35%, 0.45% and 0.55% or 0.15%, 0.22%, 0.29%, 0.36% and 0.43% from 1 to 21 or 22 to 42 days of age. Serum and tibia samples were collected for determinations of bone phosphorus retention and bone development parameters, related hormones and local bone-derived regulators of broiler chickens on d 14, 28 and 42, respectively. RESULTS: Tibia ash phosphorus, total phosphorus accumulation in tibia ash (TPTA), bone mineral concentration (BMC), bone mineral density (BMD), bone breaking strength (BBS), and ash on d 14, 28 or 42, serum 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) on d 28 and 42, mRNA expressions of tibia fibroblast growth factor 23 (FGF23) and dentin matrix protein 1 (DMP1) on d 14 and 28 increased linearly or quadratically (P < 0.05), while serum parathyroid hormone (PTH) on d 28, tibia alkaline phosphatase (ALP) on d 14, 28 and 42, bone gal protein (BGP) on d 14, and mRNA expression of tibia phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PHEX) on d 14 and 28 decreased linearly or quadratically (P < 0.04) as dietary NPP level increased. TPTA, BMC, BMD, and ash on d 28 and 42, BBS on d 28, and ash phosphorus on d 42 were positively correlated (r = 0.389 to 0.486, P < 0.03) with serum 1,25(OH)2D3. All of the above parameters were positively correlated (r = 0.380 to 0.689, P < 0.05) with tibia DMP1 mRNA expression on d 14, 28 and 42, but negatively correlated (r = - 0.609 to - 0.538, P < 0.02) with serum PTH on d 28, tibia ALP on d 14, 28 and 42, and BGP on d 14. TPTA, BMC and ash on d 14 and BMD on d 28 were negatively correlated (r = - 0.397 to - 0.362, P < 0.03) with tibia PHEX mRNA expression, and BMD on d 28 was positively correlated (r = 0.384, P = 0.04) with tibia FGF23 mRNA expression. CONCLUSIONS: These results suggested that bone phosphorus retention and bone development parameters had moderate to strong correlations with serum PTH and 1,25(OH)2D3 and tibia DMP1, PHEX, FGF23, ALP and BGP in broilers during the whole growth period, and thus they might be partly regulated by these related hormones and local bone-derived regulators.

Zinc alleviates the heat stress of primary cultured hepatocytes of broiler embryos via enhancing the antioxidant ability and attenuating the heat shock responses.

Zinc (Zn) has been shown to attenuate the adverse effects of heat stress on broilers, but the mechanisms involving this process remain unclear. We aimed to investigate possible protective mechanisms of Zn on primary cultured hepatocytes of broiler embryos subjected to heat stress. Three experiments were conducted. In Exp. 1, hepatocytes were treated with 0, 50, 100, 200, or 400 µmol/L added Zn as inorganic Zn sulfate (iZn) for 12, 24 or 48 h. In Exp. 2, cells were exposed to 40 °C (a normal temperature [NT]) and 44 °C (a high temperature [HT]) for 1, 2, 4, 6, or 8 h. In Exp. 3, cells were preincubated with 0 or 50 µmol/L Zn as iZn or organic Zn lysine chelate (oZn) for 8 h under NT, and then incubated with the same Zn treatments under NT or HT for 4 or 6 h. The biomarkers of antioxidative status and heat stress in cells were measured. The results in Exp. 1 indicated that 50 µmol/L Zn and 12 h incubation were the optimal conditions for increasing antioxidant ability of hepatocytes. In Exp. 2, the 4 or 6 h incubation under HT was effective in inducing heat shock responses of hepatocytes. In Exp. 3, HT elevated (P < 0.01) malondialdehyde content and expressions of heat shock protein 70 (HSP70) mRNA and protein, as well as HSP90 mRNA. However, Zn supplementation increased (P < 0.05) copper zinc superoxide dismutase (CuZnSOD) activity and metallothionein mRNA expression, and effectively decreased (P < 0.05) the expressions of HSP70 mRNA and protein, as well as HSP90 mRNA. Furthermore, oZn was more effective (P < 0.05) than iZn in enhancing CuZnSOD activity of hepatocytes under HT. It was concluded that Zn (especially oZn) could alleviate heat stress of broiler hepatocytes via enhancing their antioxidant ability and attenuating heat shock responses.

Organic iron absorption and expression of related transporters in the small intestine of broilers.

An experiment was conducted to investigate the effect of organic and inorganic Fe sources on Fe absorption and expression of related transporters in the small intestine of broilers. Iron-deficient intact broilers (7-day-old) were fed an Fe-unsupplemented corn-soybean meal basal diet or the basal diet supplemented with 60 mg Fe/kg as Fe sulfate (FeSO4•7H2O), Fe-Met with weak chelation strength (Fe-Met W), Fe-proteinate with moderate chelation strength (Fe-Prot M) or Fe-proteinate with extremely strong chelation strength (Fe-Prot ES) for 14 d. The plasma Fe contents were enhanced (P < 0.02) by Fe addition, and greater (P < 0.0002) in Fe-Prot M and Fe-Prot ES groups than in Fe-Met W and FeSO4 groups. Supplemental Fe decreased (P < 0.03) the divalent metal transporter 1 (DMT1) mRNA levels in the duodenum and jejunum, and ferroportin 1 (FPN1) mRNA levels in the duodenum on d 21, but no differences (P > 0.20) were detected among different Fe sources. Regardless of Fe source, the mRNA levels of DMT1 and FPN1 were higher (P < 0.02) in the duodenum than in the jejunum and ileum, and in the jejunum than in the ileum (P < 0.05). However, Fe addition did not affect (P > 0.10) the mRNA levels of amino acid transporters and protein levels of DMT1 and FPN1 in the small intestine of broilers. These results indicate that organic Fe sources with stronger chelation strength showed higher Fe absorption in broilers in vivo; the mRNA expression of Fe and amino acid transporters varied along with the extension of the small intestine; the absorption of Fe as organic Fe chelates was not mediated by the amino acid transporters in intact chicks in this study.