In ovo feeding of creatine pyruvate modulates growth performance,energy reserves and mRNA expression levels of gluconeogenesis and glycogenesis enzymes in liver of embryos and neonatal broilers

The effects of in ovo feeding (IOF) of creatine pyruvate (CrPyr) on the growth performance, energy reserves and mRNA expression levels of gluconeogenesis and glycogenesis enzymes in liver of late‐term embryos and neonatal broilers were investigated. After candling on 16 day of incubation, a total of 960 eggs were randomly assigned to three treatments: (i) non‐injected control, (ii) saline group injected with 0.6 ml of 0.75% physiological saline and (iii) Creatine pyruvate group injected with 0.6 ml of physiological saline containing 12 mg CrPyr/egg. After hatching, 120 male chicks with average body weight (BW) were randomly allocated into each treatment group for a 7‐day feeding trial. The results showed that broilers subjected to CrPyr treatment had higher BW than those of the control and saline groups on 1, 3 and 7 day post‐hatch, as well as the yolk sac weight on 19 day of incubation (19 E), the day of hatch and 3 day post‐hatch (p < .05). Compared with the control and saline groups, IOF of CrPyr increased the plasma creatine concentration on the day of hatch, and the plasma pyruvate concentration on the day of hatch and 3 day post‐hatch (p < .05). Moreover, IOF of CrPyr increased the liver pyruvate and glucose concentrations on 19 E and the day of hatch, and the liver glycogen concentration during the experiment (p < .05). Broilers in the CrPyr group showed increased mRNA expression levels of pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PEPCK) and glycogen synthase 2 (GYS2) on 19 E and the day of hatch (p < .05). These results indicated that IOF of CrPyr increased energy reserves in liver of embryos and neonatal broilers possibly through upregulating the mRNA expression levels of PC, PEPCK and GYS2, which could benefit the increase of BW in broilers on 7 day post‐hatch.     

In ovo feeding of nutrients and its impact on post‐hatching water and feed deprivation up to 48 hr,energy status and jejunal morphology of chicks using response surface models

A Box–Behnken design (BBD) in a response surface methodology (RSM) was used to investigate the response of broiler chicks to in ovo feeding (IOF) of beta‐hydroxy beta‐methylbutyrate (HMB), dextrin and the timing of the first water and feed deprivation. On day 18th of incubation, 1,500 eggs were randomly assigned to 15 experimental runs of BBD, each with 4 replicates, as 3 levels IOF of HMB (0%, 0.5% and 1%) and dextrin (0%, 20% and 40%), and 3 levels of the first water and feed deprivation (6, 27 and 48 hr). Day‐old chicks from each replicate were then used to assess the effect of IOF and time first water and feed access on chick's responses. The IOF of dextrin leads to respectively 9.7%–15.5% lower hatchability for 20% and 40% inclusion (p < .05), whereas HMB inclusion appeared with no effect on hatchability (p > .05). Administration of dextrin or HMB into the amnion of embryos elevated length, width and surface area of villus, and increased glycogen content of liver and breast (p < .05). In all parameter models, the linear terms showed highest contribution (R² = 0.81–0.97) to explain existing variation in chick's responses. The first water and feed deprivation had largest effect on BW2 and glycogen content of liver and breast. It is concluded that if possible, place chicks before 7 hr of hatch to preserve BW loss and have maximum response from IOF. If not possible, use IOF with 40% dextrin + 0.5% HMB to preserve gut integrity and energy status up to 48 hr. This should give advantage to chicks to recover fast after feeding, but that would have to be confirmed by trials growing birds to slaughter age.     

In ovo carbohydrate supplementation modulates growth and immunity‐related genes in broiler chickens

A study was undertaken to investigate the role of in ovo administrated carbohydrates on the expression pattern of growth and immune‐related genes. In ovo injections (n = 400) were carried out on the 14th day of incubation into the yolk sac/amnion of the broiler chicken embryos. Expression of growth‐related genes: chicken growth hormone (cGH), insulin‐like growth factor‐I & II (IGF‐I & II) and mucin were studied in hepatic and jejunum tissues of late‐term embryo and early post‐hatch chicks. Expression of candidate immune genes: Interleukin‐2, 6, 10 and 12 (IL‐2, IL‐6, IL‐10 and IL‐12), Tumour necrosis factor‐alpha (TNF‐α) and Interferon gamma (IFN‐γ) were studied in peripheral blood monocyte cells of in ovo‐injected and control birds following antigenic stimulation with sheep RBC (SRBC) or mitogen concanavalin A (Con‐A). Glucose injection significantly increased the expression of IGF‐II gene during embryonic period and both cGH and IGF‐II in early post‐hatch period, while ribose‐injected chicks had higher expression of IGF‐II gene during embryonic stage. Enhanced mucin gene expression was also observed in fructose‐injected chicks during embryonic age. Glucose‐injected chicks had higher expression of IL‐6 or IL‐10, while those injected with fructose or ribose had higher expression of IL‐2, IL‐12 and IFN gamma. It is concluded that in ovo supplementation of carbohydrates might help in improving the growth of late‐term embryos and chicks. In ovo glucose could modulate humoral‐related immunity, while fructose or ribose might help in improving the cellular immunity in broiler chickens.     

In ovo L‐arginine supplementation stimulates myoblast differentiation but negatively affects muscle development of broiler chicken after hatching

In this study, we tested the hypothesis that in ovo feeding (IOF) of L‐arginine (L‐Arg) enhances nitric oxide (NO) production, stimulates the process of myogenesis, and regulates post‐hatching muscle growth. Different doses of L‐Arg were injected into the amnion of chicken embryos at embryonic day (ED) 16. After hatching, the body weight of individual male chickens was recorded weekly for 3 weeks. During in vitro experiments, myoblasts of the pectoralis major (PM) were extracted at ED16 and were incubated in medium containing 0.01 mm L‐Arg, 0.05 mm L‐Arg, and (or) 0.05 mm L‐nitro‐arginine‐methyl‐ester (L‐NAME), an inhibitor of nitric oxide synthase (NOS). When 25 mg/kg L‐Arg/initial egg weight was injected, no difference was observed in body weight at hatch, but a significant decrease was found during the following 3 weeks compared to that of the non‐injected and saline‐injected control, and this also affected the growth of muscle mass. L‐NAME inhibited gene expression of myogenic differentiation antigen (MyoD), myogenin, NOS, and follistatin, decreased the cell viability, and increased myostatin (MSTN) gene expression. 0.05 mm L‐Arg stimulated myogenin gene expression but also depressed muscle cell viability. L‐NAME blocked the effect of 0.05 mm L‐Arg on myogenin mRNA levels when co‐incubated with 0.05 mm L‐Arg. L‐Arg treatments had no significant influence on NOS mRNA gene expression, but had inhibiting effect on follistatin gene expression, while L‐NAME treatments had effects on both. These results suggested that L‐Arg stimulated myoblast differentiation, but the limited number of myoblasts would form less myotubes and then less myofibers, while the latter limited the growth of muscle mass.     

Effects of delaying post‐hatch feeding on lipid peroxidation and antioxidant enzyme mRNA expression in the pectoralis major muscle of newly hatched chicks

Excessive lipid peroxidation negatively affects the physiological response and meat quality of chickens. Delaying post‐hatch feeding was previously found to increase lipid peroxidation in the skeletal muscle of finishing broiler chickens. The aims of this study were to investigate the effects of delayed post‐hatch feeding on lipid peroxidation and the mRNA expressions of antioxidant enzymes in the pectoralis major muscle of broiler chicks during the post‐hatching period. Newly hatched chicks either had immediate free access to feed (freely‐fed chicks) or had no access to feed from 0 to 2 days old (delayed‐fed chicks), after which both groups were fed ad libitum until 4 or 13 days old. The lipid peroxidation level was higher in the delayed‐fed than freely‐fed chicks at 2, 4, and 13 days old. At 2 days old, the mRNA expressions of Cu/Zn‐SOD, Mn‐SOD, and GPX7 were lower in the delayed‐fed than freely‐fed chicks, while catalase mRNA levels did not differ. Furthermore, at 4 and 13 days old, lower mRNA expressions of Cu/Zn‐SOD and Mn‐SOD were observed in the delayed‐fed than freely‐fed chicks. These results suggest that delaying post‐hatch feeding reduces the mRNA levels of Cu/Zn‐SOD and Mn‐SOD, consequently affecting muscle lipid peroxidation in chicks during subsequent growth.     

Effects of in ovo injection of chrysin,quercetin and ascorbic acid on hatchability,somatic attributes,hepatic oxidative status and early post‐hatch performance of broiler chicks

An experiment was conducted to evaluate the effects of in ovo injection of chrysin, quercetin and ascorbic acid on hatchability, somatic attributes, hepatic antioxidant status and early post‐hatch growth performance of broiler chicks. Four hundred and eighty embryonated broiler breeder eggs containing live 18‐day‐old embryos were divided into six groups of 80 eggs each. One group remained intact and served as a control group (i), whereas the other five groups were injected with the prepared injection solutions as follows: (ii) 0.05 ml distilled water; (iii) 0.05 ml distilled water containing 6 mg ascorbic acid; (iv) 0.05 ml dimethyl sulfoxide (DMSO); (v) 0.05 ml DMSO containing 4.5 mg quercetin; and (vi) 0.05 ml DMSO containing 4.5 mg chrysin. The hatchability rate, hatching weight, residual yolk sac weight, yolk sac‐free body weight, liver weight, hepatic glutathione peroxidase and total superoxide dismutase activities, as well as malondialdehyde concentrations, were not affected by the injected solutions. There were no differences between chicks hatched from the control and in ovo injected eggs in weight gain, feed intake and feed conversion ratio from 0 to 11 days of age. However, the specific contrast performed between the in ovo injected groups and intact eggs revealed that in ovo injection significantly increased hatchability rate (p = .0493). This finding also implies that our injection procedure was harmless. In conclusion, the intra‐egg injection of chrysin, quercetin or ascorbic acid at the injection rates used in this study did not have a significant effect on hatchability, somatic characteristics, early growth performance and hepatic antioxidant status of broiler chicks. However, the overall hatchability was higher in the in ovo injected eggs as compared to non‐injected ones. These findings also confirmed the harmlessness of the procedure developed for in ovo injection in this study.     

Tibiotarsus bone characteristics and tibial dyschondroplasia incidence of broilers fed diets supplemented with leucine and valine

Two experiments were conducted to study the effect of standardized ileal digestible (SID) leucine and valine levels on tibiotarsus bone characteristics and the incidence of tibial dyschondroplasia of broilers from day 1 to 21 (Experiment I) and day 21 to 42 post‐hatch (Experiment II). Each experimental phase was evaluated independently. In both experiments, a total of 1,500 one‐day‐old Cobb 500 male broiler chickens were distributed in a completely randomized design 5 × 5 factorial arrangement for a total of 25 treatments. The SID leucine and valine levels were ranged from 10.0 to 19.6 g/kg, and 6.0 to 12.0 g/kg from day 1 to 21 post‐hatch, respectively, while day 21 to 42 post‐hatch ranged from 10.0 to 18.0 g leucine/kg, and 5.2 to 11.2 g valine/kg. Serum calcium and phosphorus, bone concentrations of calcium, phosphorus and ash, diameter and Seedor index of the tibiotarsus were not affected (p > .05) by the treatments at 21 or 42 days of age. There was an interaction (p ≤.06) between the SID levels of leucine and valine on tibiotarsus breaking strength at 21 days, but not at 42 days of age (p > .05). Tibiotarsus breaking strength was maximized in broilers from day 1 to 21 with the dietary levels of leucine and valine at 14.2 and 9.0 g/kg respectively. Dietary leucine levels reduced linearly (p < .05) the hypertrophic zone of tibiotarsus cartilage at 21 days of age. Therefore, leucine and valine supplementation interact positively on bone strength of broilers from day 1 to 21 post‐hatch. Leucine can be a useful amino acid for reducing the hypertrophic cartilage zone in broilers from day 1 to 21, but not from day 21 to 42 post‐hatch.     

Glycerol in ovo feeding as an energy substrate improves performance of broilers from young breeders

Glycerol is one of the substrates used for glycogen production by the chicken embryo, which is the predominant energy source during the last days of incubation and during hatching. The objective of the present study was to evaluate the in ovo feeding (IOF) of glycerol in the light and heavy broiler eggs derived from breeders of two different ages. Two experiments, with 672 eggs each, were carried out. The only difference between the experiments was breeder age: 32 weeks old in Exp. I and 60 weeks old in Exp. II. A completely randomized experimental design in a 3 × 2 factorial arrangement was applied. Treatments consisted of three glycerol IOF doses (0, 6, or 12 mg/ml) and two egg weights (light or heavy). Incubation parameters, glycogen reserves and live performance parameters (1–7 days of age) were evaluated. Hatch of fertile eggs, embryo mortality after IOF and the number of early‐hatching chicks were not affected by the treatments in both experiments. Hatchlings from heavy eggs (68.03 ± 0.64 g) laid by young breeders and receiving 6 mg glycerol/ml showed higher liver glycogen levels than those injected with 0 or 12 mg/ml. Glycerol IOF of embryos from young breeders increased feed intake and weight gain at 7 days of age, independently of egg weight. However, different glycerol dosages had no effect on the performance of the progeny of 60‐week‐old breeders. These results show that glycerol may be used as an IOF ingredient without affecting incubation parameters. The chickens from young breeders had greater glycogen deposition with inoculation of 6 mg/ml of glycerol and better performance with glycerol administration. However, glycerol IOF did not improve the performance of the progeny of 60‐week‐old breeders. Therefore, glycogen IOF may be recommended for eggs laid by young breeders.     

In ovo injection of branched‐chain amino acids: Embryonic development,hatchability and hatching quality of turkey poults

In this study, the influence of a branched‐chain amino acid blend (BCAA composed of 3 l ‐leucine:1 l ‐valine:2 l ‐isoleucine) injected into the amniotic fluid was evaluated for embryonic growth, yolk‐sac (YS) utilization and development of gastrointestinal tract (GIT) and skeletal muscles of turkey embryos from day 24 of incubation (24E) to hatching, together with hatchability, poult quality and liver L* (lightness), a* (redness) and b* (yellowness) values at hatch. At day 22 of incubation, embryonated eggs (n = 240) were assigned to three treatments, that is, eggs were not injected (control, NC) or injected with 1.5 ml sterile solution with 0.9% salt (SA) or 0.2% BCAA blend (BCAAb). These solutions were injected manually into the amniotic fluid of the embryonated eggs. To determine weights and lengths (where appropriate) of the studied organs and tissues, four embryonated eggs and poults per treatment were selected at 24E and at hatch. While the BCAAb decreased the YS and embryo weight, hatchability and the liver L* value, it increased the weight and quality of poults and the weights of breast and thigh muscles at hatch. In conclusion, the in ovo feeding of the BCAA blend negatively affected hatchability but positively affected hatching weight and poult quality by improving development of skeletal muscles and by regulating energy metabolism.     

In ovo injection of flavanone on bone quality characteristics,biochemical parameters and antioxidant enzyme status of blood in daily chicks

In ovo injection (IOI) of Naringin (N), flavanone was examined on post‐hatch blood biochemical parameters, antioxidant status and bone characteristics. Fertile eggs (n = 700) were distributed in seven groups with 100 eggs. On 14th and 17.5th days of incubation, four groups were injected using 15 or 30 mg active ingredient levels of naringin/0.5 ml saline/egg, low and high level, into amnion sac. Controls include sham (injected normal saline, 0.5 ml/egg on day 14 and 17.5th) and un‐injected group. IOI of high naringin and saline on 14th day of incubation resulted in lower hatchability and then higher mortality in last week of embryonic life. On day hatch, high levels of injected groups more body weight compared to the control. Chick length was increased at high levels of naringin on day 17.5th compared to control and saline injected. Quality traits of bones were improved in naringin‐injected groups compared to control. IOI of naringin influenced thyroid hormones on 14th day of incubation. Naringin groups influenced the Alkaline phosphatase (ALP), Calcium (Ca), superoxide dismutase (SOD), blood biochemical and lipids. Totally, amniotic IOI of naringin in last days of developing embryo may be useful for hatched chick, development of leg long bone or effect on biochemical metabolites by levels of flavanone that it needs more research.     

In ovo feeding of carbohydrates for broilers—a systematic review

The purpose of this systematic review was to evaluate the evidence that the injection of carbohydrate‐based solutions into embryonated eggs improves broiler performance. A literature search was conducted in April 2017 using the keywords broiler, carbohydrate, in ovo, nutrition and poultry. Only papers that involved in ovo carbohydrate injections in poultry were used in this study. After specific selection criteria, 17 papers were selected. The quality scoring system of the selected studies was based on the injection methodology, use of control groups, type of solution injected, period of injection, egg and hens characteristics, number of variables analysed and the statistical design. Among papers, there was no standardised procedure in to inoculate the solutions. Nevertheless, in general, in ovo feeding of carbohydrates decreases the hatch rate, improves the hatch weight, but it does not seem to influence the post‐hatch performance of broilers. The inoculation of 75 mg of glucose in the albumen seems to bring better results. Further studies are needed to improve the technical methodology of in ovo injections for commercial use.     

The interaction between maternal and post‐hatch n‐3 fatty acid supplementation in broiler diets

This study investigated whether offspring from n‐3‐supplemented breeders have an enhanced performance and immune organ weight when fed a post‐hatch n‐3‐enriched diet in comparison with their control‐fed counterparts and the importance of timing of omega‐3 supplementation. Therefore, 480 Ross‐308 broiler breeder hens were fed one of four different diets (120/treatment). The control diet (CON) was a basal diet, rich in n‐6 fatty acids (FA). The three other diets were enriched in n‐3 FA, formulated to obtain a different EPA/DHA ratio of 1/1 (EPA = DHA), 1/2 (DHA) or 2/1 (EPA). At 33 weeks of age, eggs were incubated to obtain 1440 offspring. They were set up according to their maternal diet and sex in 48 pens of 30 chicks each (12 pens per maternal treatment: six male and six female). Half of the offspring were given a post‐hatch control diet, whereas to other half received an n‐3‐supplemented diet. Zootechnical performance was followed for starter, grower and finisher phase, and at the end of each phase two, chicks per pen were sacrificed to determine the weight of the immune organs. No interaction was found between maternal and post‐hatch n‐3 treatment for zootechnical performance. An interaction arose between the maternal and post‐hatch n‐3 supplementation for proportional bursa weight at day 1 and day 14 and proportional liver weight at day 14, but effects on immune organ weight were rather limited. Offspring post‐hatch n‐3 supplementation did not enhance maternal n‐3 supplementation.     

Dominant follicles development and estradiol‐17β concentrations in non‐ovulating and ovulating post‐partum Bulgarian Murrah buffaloes

This study was designed to evaluate the dominant follicles development and the estradiol‐17β concentrations in non‐ovulating and ovulating post‐partum buffaloes. Sixteen Bulgarian Murrah buffaloes were submitted to transrectal ultrasonographic examination from the 1st post‐partum day until day 50, 3 days apart. The follicular diameter of the different categories of follicles and the ovulations was recorded. The animals were allocated into two groups: I (n = 6) non‐ovulating and II (n = 10) ovulating buffaloes. Serum estradiol‐17β concentrations on the days for dominant follicle registration were measured by enzyme‐linked immunosorbent assay. The results were statistically processed by analysis of variance, non‐parametric and correlation analysis. The mean intervals between calving and first dominant follicle detection differed significantly (p < .05) among the groups (19.5 ± 6.2 vs. 13.8 ± 5.1 days), while the mean intervals between registered dominant follicles from two successive waves were comparable. The mean follicular diameters for the same category follicles in both groups were similar. Different estradiol‐17β concentrations (p < .05) for the first dominant follicle between non‐ovulating (23.5 ± 7.0 pg/ml) and ovulating (33.3 ± 8.4 pg/ml) buffaloes were determined. The cumulative percentages of buffaloes with firstly detected dominant follicle and ovulating animals correlated positively (r ≥ .84; p < .05) to post‐partum days. In conclusion, non‐ovulating and ovulating post‐partum Bulgarian Murrah buffaloes showed differences in the development of the first dominant follicle and estradiol‐17β concentrations during the time of dominant follicles detection.     

The effect of Epigallocatechin‐3‐gallate on small intestinal morphology,antioxidant capacity and anti‐inflammatory effect in heat‐stressed broilers

This study was to investigate the effects of Epigallocatechin‐3‐gallate (EGCG) on intestinal morphology, antioxidant capacity and anti‐inflammatory response in heat‐stressed broiler. A total of 192 2‐week‐old Arbour Acres broilers chickens were divided into four groups with six replicates per group and eight chickens per replicate: one thermoneutral control group (28°C, group TN), which was fed the basal diet; and three cyclic high‐temperature groups (35°C from 7:00 to 19:00 hr; 28°C from 19:00 hr to 7:00 hr, heat stress group), which were fed the basal diet supplementation with EGCG 0 mg/kg (group HS0), 300 mg/kg (group HS300) and 600 mg/kg (group HS600). The gut morphology and intestinal mucosal oxidative stress indicators, as well as intestinal barrier‐related gene expression, were analysed. The results showed that compared with group TN, heat stress reduced the villus height (VH), activities of glutathione peroxidase (GSH‐Px), superoxide dismutase (SOD)and catalase (CAT), increased the crypt depth (CD) and malondialdehyde (MDA)content at 21, 28 and 35 days (p < 0.05). After the heat‐stressed broilers were supplemented with EGCG, VH, VH/CD (V/C), and the activities of GSH‐Px, SOD and CAT were increased, and CD and MDA content were reduced compared with those in group HS0 without EGCG supplementation at 21, 28 and 35 days (p < 0.05). The EGCG supplementation promoted the gene expression of nuclear factor‐erythroid 2‐related factor 2 (Nrf2), Claudin‐1, Mucin 2 (Muc2) and alleviated the nuclear factor‐kappa B (NF‐κB) and lipopolysaccharide‐induced tumour necrosis factor (LITAF) gene expression compared with group HS0 (p < 0.05). Moreover, intestinal morphology was strongly correlated with antioxidant ability and inflammatory response. In conclusion, EGCG alleviated the gut oxidative injury of heat‐stressed broilers by enhancing antioxidant capacity and inhibiting inflammatory response.     

Effects of dietary inclusion of silymarin on performance,intestinal morphology and ileal bacterial count in aflatoxin‐challenged broiler chicks

This study was conducted to investigate the effect of dietary supplementation of silymarin on performance, jejunal morphology and ileal bacterial population in broiler chicks intoxicated with a mix of aflatoxins. A total of three hundred thirty six 7‐day‐old Ross broiler chicks were randomly distributed between seven experimental groups with four replicates of 12 birds each. Experimental treatments consisted of a control group (unchallenged), and a 2 × 3 factorial arrangement, including two aflatoxin levels (0.5 and 2 ppm) and three levels of silymarin (0, 500 and 1000 ppm). Birds were challenged with a mix of aflatoxins from 7 to 28 days of age. Results showed that increasing aflatoxin level resulted in decreased average daily feed intake (ADFI) and weight gain (ADWG), consequently impaired feed conversion ratio (FCR) throughout the trial period. Dietary supplementation of silymarin resulted in the marked increases in ADFI and ADWG, and improved FCR values in aflatoxin‐challenged chicks. Ileal bacterial populations at days 28 and 42 of age were increased by incremental levels of aflatoxins. On the other hand, dietary silymarin supplementation suppressed ileal populations of Escherichia coli, Salmonella, Klebsiella and total negative bacteria in aflatoxicated birds. Increase in dietary aflatoxin level resulted in the decreased villi height, villi height‐to‐crypt depth ratio (VH:CD), villi surface area and apparent villi absorptive area, while it increased crypt depth, goblet cell count and lymphoid follicular diameter. Feeding silymarin at the level of 1000 ppm increased villi height and VH:CD in aflatoxicated birds. Present results indicate that dietary inclusion of silymarin could improve performance by suppressing ileal bacteria and enhancing absorptive surface area in aflatoxin‐challenged broiler chicks.     

Effect of laboratory‐isolated Lactobacillus plantarum LGFCP4 from gastrointestinal tract of guinea fowl on growth performance,carcass traits,intestinal histomorphometry and gastrointestinal microflora population in broiler chicken

The study aimed to investigate the effect of feed supplements, viz Lactobacillus plantarum LGFCP4 (laboratory isolate from GIT of Guinea fowl), Lactobacillus acidophilus (NCDC, Karnal) and in‐feed antibiotic bacitracin methylene disalicylate (BMD) on growth performance, FCR, carcass traits and immune organs weight, intestinal histomorphometry and gastrointestinal microflora population in broiler chickens. In a completely randomized design, CARIBRO‐Dhanraja broiler chicks (n = 160) were used with four treatment groups. During the entire experimental duration of 35 days, treatment groups were provided with different dietary treatments (T1 – basal diet (negative control), T2 – antibiotic growth promoter BMD 20 g/100 kg feed (positive control), T3 – 1 × 10⁸ cfu of L. acidophilus/gm‐fermented feed +MOS 1 g/kg feed and T4 – 1 × 10⁸ cfu of laboratory‐isolated L. plantarum LGFCP4/gm‐fermented feed+ MOS 1 g/kg feed. After 35 days of experimental period, no significant results have been observed in different growth performance traits among treatment groups. Cut‐up parts and edible organs' weight remained unaffected by dietary supplementation, whereas weight of immune organs were significantly higher (p < 0.05) in L. plantarum LGFCP4‐supplemented group. At the end of feeding trial, significantly (p < 0.05) lower E. coli count was observed in crop of T4 birds, while in ileum, T2 and T3 showed lower count. In caeca, T2 group showed lowest E. coli count. Salmonella count in crop and ileum was significantly (p < 0.05) low in T3 and T4, while in caeca, T2 group showed lowest count. In terms of histomorphometry, duodenal villous height (VH), crypt depth (CD) and VH:CD ratio were higher for T3 and T4 and lowest values were obtained for T2 group. The results of the study showed that L. plantarum LGFCP4 isolated from GIT of guinea fowl can effectively replace in‐feed antibiotic growth promoters in broiler diets by altering intestinal villi morphology and improving the gut health by reducing the pathogenic microbial load.     

Study on differentiation during embryonic development across selective and ancestral breeds

In order to explore the effect of strain on diverging post‐hatch muscle properties, muscle regulation during embryo development was investigated in selected and unselected breeds. Four broiler strains were used: JingNing (JN) chicken (a Chinese native chicken), HuangYu (HY) broiler, BaiYu (BY) broiler and Hyline layer (commercial crossbred chickens). Results showed that the four breeds had almost the same characteristic during different incubation periods. BY broilers moved more than JN and Hyline layers from Hamburger & Hamilton stage (HH)24 to HH31 (P < 0.05). HY broilers moved more than JN and Hyline layers from HH27 to HH31 (P < 0.01). All the embryos were heavier daily from HH24 to ED18 (P < 0.05); broilers presented greater body weights than JN and hyline layers (P > 0.05); broilers presented smaller fiber diameter than JN chickens before HH31 (P > 0.05). From then on, JN chicken exhibited smaller fiber diameter compared to the broilers (P > 0.05). Western blotting indicated all the breeds had continuous insulin‐like growth factor‐I (IGF‐I) expression, with the highest expression level in broilers from HH19 to HH24 and highest expression level in JN chicks from HH27 to HH31. The results indicated that the diverging growth among breeds was already shown in embryonic stages; the different expression patterns of IGF‐I may be involved in cell proliferation and differentiation.     

The effect of n‐6/n‐3 fatty acid ratios on broiler breeder performance,hatchability, fatty acid profile and reproduction

This experiment was conducted to study the effect of dietary omega6 (n‐6) to omega3 (n‐3) fatty acid (FA) ratios on performance and reproduction of broiler breeders. In experiment 1, 400 females and 40 males (30 week age) of Ross 308 broiler breeder (20 females and two males in each pen) were randomly assigned to one of the four diets with n‐6/n‐3 FA ratios of 4, 6, 8 and 16 (control). As a measure of hatchability, fertility of eggs and general incubation traits, 1,200 eggs (60 eggs from each pen) were collected and incubated for 21 days and embryo liver and brain fatty acid profile in 14 and 21 days were determined. In experiment 2, 48 males (three males in each pen) randomly assigned to one of the four diets with n‐6/n‐3 FA ratios of 4, 6, 8 and 16 (control). Semen was collected twice weekly, and semen volume, spermatozoa concentration and motility and alive and dead spermatozoa were estimated. Egg production and egg mass were decreased by n‐6/n‐3 FA ratios of 4:1 and 6:1 (p < .05). There were no significant differences between treatments on breeder's body weight, eggs fertility and hatchability, embryonic mortality and semen features. Linolenic acid, eicosapentaenoic acid, docosahexaenoic acid and total n‐3 of egg yolk, semen, testis and liver and brain of embryo and day‐old chicken were increased while concentration of linoleic acid, arachidonic acid and docosatetraenoic acid of mentioned tissues were decreased by increasing n‐6/n‐3 FA ratios (p > .05). In conclusion, absolute amount of n‐3 and n‐6 FAs in broiler breeder diet may be more important than n‐6/n‐3 FA ratios and to consider reproductive and performance traits of breeders, it is necessary to supply higher levels of n‐3 and n‐6 FA with respect to n‐6/n‐3 FA ratios.     

Developmental regulation and modulation of apoptotic genes expression in sheep oocytes and embryos cultured in vitro with L‐carnitine

The objective of this study was to find out the impact of L‐carnitine (10 mM) on developmental regulation of preimplantation sheep embryos cultured in vitro when supplemented in maturation medium and post‐fertilization medium separately. Subsequent objective was to observe the L‐carnitine‐mediated alteration in expression of apoptotic genes (Bcl2, Bax, Casp3 and PCNA) in sheep oocytes and developing embryos produced in vitro. Oocytes matured with L‐carnitine showed significantly (p < .05) higher cleavage (67.23% vs 43.12%), morula (47.65% vs 28.58%) and blastocysts (32.12% vs 13.24%) percentage as compared to presumptive zygotes cultured with L‐carnitine during post‐fertilization period. So it is suggested to use L‐carnitine during maturation than post‐fertilization period. Antiapoptotic and proliferative effects of L‐carnitine were confirmed by inducing culture medium with actinomycin D (apoptotic agent) and TNFα (antiproliferative agent), respectively, with and without L‐carnitine. Oocytes and embryos cultured with actinomycin D and TNFα showed developmental arrest with significant (p < .05) decrease in morula and blastocysts percentage but s upplementation of L‐carnitine to actinomycin D and TNFα induced culture medium showed similar result as that of control . L‐carnitine supplementation during IVM significantly (p < .05) upregulated the expression of Bcl2 and PCNA genes in majority of the developmental stages. Although L‐carnitine upregulated the expression of Bax in initial developmental stages but downregulated at latter part, whereas the expression of Casp3 was upregulated upto 16‐cell stage but after that there was no difference in expression. Expression of GAPDH gene was not affected by L‐carnitine supplementation. In conclusion, L‐carnitine acted as an antiapoptotic and proliferative compound during embryo development and supplementation of L‐carnitine during IVM altered the expression of apoptotic genes in the developmental stages of embryos.     

Effect of different penetrating and non‐penetrating cryoprotectants and media temperature on the cryosurvival of vitrified in vitro produced porcine blastocysts

The aim of this study was to determine the most efficient vitrification protocol for the cryopreservation of day 7 in vitro produced (IVP) porcine blastocysts. The post‐warm survival rate of blastocysts vitrified in control (17% dimethyl sulfoxide + 17% ethylene glycol [EG] + 0.4 mol/L sucrose) and commercial media did not differ, nor did the post‐warm survival rate of blastocysts vitrified in medium containing 1,2‐propandiol in place of EG. However, vitrifying embryos in EG alone decreased the cryosurvival rate (55.6% and 33.6%, respectively, p < .05). Furthermore, the post‐warm survival rates of blastocysts vitrified with either trehalose or sucrose as the non‐penetrating cryoprotectant did not differ. There was also no significant difference in post‐warm survival of blastocysts vitrified in control (38°C) media and room temperature (22°C) media with extended equilibration times, although when blastocysts were vitrified using control media at room temperature, the post‐warm survival rate increased (56.8%, 57.3%, 72.5%, respectively, p < .05). The findings show that most cryoprotectant combinations examined proved equally effective at supporting the post‐warm survival of IVP porcine blastocysts. The improved post‐warm survival rate of blastocysts vitrified using media held at room temperature suggests that the cryoprotectant toxicity exerted in 22°C media was reduced.