Slaughter performance and meat quality of Milanino chickens reared according to a specific free-range program.

The study aimed to characterize meat quality traits of Milanino chickens reared according to a specific free-range farming program. A total of 120 birds was reared straight-run in outdoor pens (8 m2/bird) from 35 d of life and fed ad libitum a low (16%) protein diet. At 180 d of age, 20 birds (10 birds/sex) were slaughtered, and carcass weight data were recorded. After processing, carcasses were refrigerated at 4°C for 24 hours. Then, the right breast and thigh with skin were collected and color parameters, pH, water-holding capacity (WHC), and chemical composition were determined. The left breast and thigh were stored at -20°C until cooking loss and tenderness evaluation. Milanino was confirmed to be a heavy breed with a sexual dimorphism in relation to adult body weight. A high general carcass yield was recorded. Milanino meat was characterized by high protein and low fat contents compared with the standard broiler meat. Differences in meat composition were recorded according to the sex: females presented higher values of dry matter (breast and thigh), protein (breast), and fat (breast and thigh) contents. The meat with skin presented an intense luminosity, and this trait was higher in the females. The muscle color was characterized by high redness and yellowness indices with differences according to the sex: Higher yellowness index was observed in female carcasses, while higher redness index was detected in male breast samples. The pH muscle values were similar to those reported in other autochthonous breeds. WHC values did not show variation between sexes. In contrast, cooking loss values recorded in thigh samples were lower in males compared to females. The degree of tenderness of Milanino meat was not affected by the sex. However, the potential loss of water and the toughness in Milanino meat were low compared to other local chicken breed meat. The present results support the breeding of Milanino chickens for meat production according to its specific straight-run free-range system.

Spermatozoa DNA and plasma membrane integrity after pellet optimized processing for cryopreservation in meat type chicken breeders.

1. Aim of this study was the development of an optimised cryopreservation pellet procedure for chicken semen and the assessment of DNA and membrane integrity in frozen/thawed spermatozoa in a Hubbard F15 meat type selected strain. 2. The following semen processing conditions were studied: spermatozoa working concentration (SWC), 1.5 vs 2 × 109 cells/ml in pre-freezing extender; equilibration of diluted semen at 5°C, 20 vs 40 min; dimethylacetamide concentration, 6% vs 9%; dimethylacetamide equilibration time at 5°C, 1 vs 30 min; thawing at 60°C for 10 vs 50°C for 30 sec. Spermatozoa viability (EtBr exclusion procedure - stress test), mobility (Accudenz® swim-down test) and subjective motility were assessed in fresh and frozen-thawed semen. 3. The lower SWC (1.5 × 109 cells/ml) and the higher dimethylacetamide concentration (9%) had positive significant effects on the recovery rate of motile (22% vs 16%) and viable spermatozoa (39 vs 34%), respectively. 4.Membrane (SYBR14-PI staining) and DNA integrity (comet assay) were assessed before and after freezing/thawing according to the optimised protocol. 5. Recovery rates of spermatozoa with undamaged plasma membrane and DNA were 41% and 76%, respectively. The distribution of spermatozoa in classes of DNA damage was also analysed and discussed. 6. It was concluded that pellet cryopreservation was a damaging process mainly for plasma membrane rather than nuclear DNA in chicken spermatozoa.

Effect of cooling rate on the survival of cryopreserved rooster sperm: Comparison of different distances in the vapor above the surface of the liquid nitrogen.

The aim of the present trial was to study the effect of different freezing rates on the survival of cryopreserved rooster semen packaged in straws. Slow and fast freezing rates were obtained keeping straws at different distances in the vapor above the surface of the nitrogen during freezing. Adult Lohmann roosters (n=27) were used. Two experiments were conducted. In Experiment 1, semen was packaged in straws and frozen comparing the distances of 1, 3 and 5cm in nitrogen vapor above the surface of the liquid nitrogen. In Experiment 2, the distances of 3, 7 and 10cm above the surfaces of the liquid nitrogen were compared. Sperm viability, motility and progressive motility and the kinetic variables were assessed in fresh and cryopreserved semen samples. The recovery rates after freezing/thawing were also calculated. In Experiment 1, there were no significant differences among treatments for all semen quality variables. In Experiment 2, the percentage of viable (46%) and motile (22%) sperm in cryopreserved semen was greater when semen was placed 3cm compared with 7 and 10cm in the vapor above the surface of the liquid nitrogen. The recovery rate of progressive motile sperm after thawing was also greater when semen was stored 3cm in the vapor above the surface of the liquid nitrogen. More rapid freezing rates are required to improve the survival of rooster sperm after cryopreservation and a range of distances from 1 to 5cm in nitrogen vapor above the surface of the liquid nitrogen is recommended for optimal sperm viability.

Growth performance, carcass characteristics and meat composition of Milanino chickens fed on diets with different protein concentrations.

Milanino is a heavy Italian chicken breed included in a conservation project of the University of Milan and is an important genetic resource for alternative production systems. This research was aimed to study the effect of the dietary protein concentration on growth, slaughter performance and meat composition in free-range reared Milanino chickens. A total of 120 Milanino chickens were fed on different protein concentrations (HP = 20% CP and LP = 16% CP), reared according to a free-range system and slaughtered at 150 and 180 d of age. Growth, slaughter performance and meat (breast and thigh) composition were recorded. The protein concentration of the diet did not affect the overall Milanino mean body weight recorded in the straight-run group in the whole rearing period. However, the growth rate within sex was significantly different between the dietary treatments: heavier females were found in the HP group from 125 d onwards, while no differences were recorded in male body weights. The protein concentration of the diet did not affect carcass weight data or meat composition. The present results suggest the use of a low-protein diet for rearing straight-run Milanino chickens for long rearing periods. However, in females, a high-protein diet is recommended from 125 d of age onwards.

Cryopreserving turkey semen in straws and nitrogen vapour using DMSO or DMA: effects of cryoprotectant concentration, freezing rate and thawing rate on post-thaw semen quality.

1. This study was designed to identify a suitable protocol for freezing turkey semen in straws exposed to nitrogen vapour by examining the effects of dimethylacetamide (DMA) or dimethylsulfoxide (DMSO) as cryoprotectant (CPA), CPA concentration, freezing rate and thawing rate on in vitro post-thaw semen quality. 2. Pooled semen samples were diluted 1:1 (v:v) with a freezing extender composed of Tselutin diluent containing DMA or DMSO to give final concentrations of 8% or 18% DMA and 4% or 10% DMSO. The semen was packaged in 0.25 ml plastic straws and frozen at different heights above the liquid nitrogen (LN2) surface (1, 5 and 10 cm) for 10 min. Semen samples were thawed at 4°C for 5 min or at 50°C for 10 s. After thawing, sperm motility, viability and osmotic tolerance were determined. 3. Cryosurvival of turkey sperm was affected by DMSO concentration. Freezing rate affected the motility of sperm cryopreserved using both CPAs, while thawing rates showed an effect on the motility of sperm cryopreserved using DMA and on the viability of sperm cryopreserved using DMSO. Significant interactions between freezing rate × thawing rate on sperm viability in the DMA protocol were found. 4. The most effective freezing protocol was the use of 18% DMA or 10% DMSO with freezing 10 cm above the LN2 surface and a thawing temperature of 50°C. An efficient protocol for turkey semen would improve prospects for sperm cryobanks and the commercial use of frozen turkey semen.

Quantitative trait loci mapping for conjugated linoleic acid, vaccenic acid and ∆(9) -desaturase in Italian Brown Swiss dairy cattle using selective DNA pooling.

A selective DNA pooling approach was applied to identify QTL for conjugated linoleic acid (CLA), vaccenic acid (VA) and Δ(9) -desaturase (D9D) milk content in Italian Brown Swiss dairy cattle. Milk samples from 60 animals with higher values (after correction for environmental factors) and 60 animals with lower values for each of these traits from each of five half-sib families were pooled separately. The pools were genotyped using the Illumina BovineSNP50 BeadChip. Sire allele frequencies were compared between high and low tails at the sire and marker level for SNPs for which the sires were heterozygous. An r procedure was implemented to perform data analysis in a selective DNA pooling design. A correction for multiple tests was applied using the proportion of false positives among all test results. BTA 19 showed the largest number of markers in association with CLA. Associations between SNPs and the VA and Δ(9) -desaturase traits were found on several chromosomes. A bioinformatics survey identified genes with an important role in pathways for milk fat and fatty acids metabolism within 1 Mb of SNP markers associated with fatty acids contents.

The post-thaw irradiation of avian spermatozoa with He-Ne laser differently affects chicken, pheasant and turkey sperm quality.

The effects of post-thaw Helium-Neon (He-Ne) laser irradiation on mobility and functional integrity of frozen/thawed chicken, pheasant and turkey spermatozoa were investigated. Cytochrome C oxidase (COX) activity was also determined as a measure of the effect of irradiation on mitochondrial bioenergetics. Semen samples from each species were collected, processed and frozen according to the pellet procedure. After thawing, each semen sample was divided into two subsamples: the first one was the control; the second one was irradiated with a single mode continuous He-Ne laser wave (wavelength 632.8 nm; 6 mW; 3.96 J/cm(2)). Then the samples were assessed for sperm mobility (Accudenz(®) swim-down test), viability (SYBR-14/PI staining), osmotic-resistance (HOS test) and COX activity. The irradiation was effective P<0.05 increasing sperm motility in the turkey semen (0.228 ± 0.01 compared with 0.294 ± 0.02). The irradiation also caused an increase (P<0.05) of the COX activity in pheasant (+135 ± 4%) and turkey (+116 ± 4%) sperm, without affecting viability and osmotic-resistance. The COX was positively correlated (P<0.05) with the viability of chicken sperm, however no significant interactions were found between mobility and COX activity in the three avian species. Due to the difference in energetic metabolism among avian species used in this study, the He-Ne laser irradiation has a differential action on bio-stimulation of turkey, chicken and pheasant spermatozoa. The present results are the first to elucidate the possibility for restoration of motility of cryopreserved avian spermatozoa by bio-stimulation provided via He-Ne laser irradiation.

Egg related parameters affecting fertility and hatchability in the Italian bantam breed Mericanel della Brianza.

Local chicken breeds are a vital reservoir of gene resources and their conservation has a technical role related to the future development of the productive system, as well as a social-cultural role. The aim of this study was to evaluate the effects of egg weight, egg storage period and egg weight loss on hatchability of fertile eggs in the Italian bantam breed Mericanel della Brianza. Fourteen females and eight males were kept in floor pens and divided in 8 families (1M:1 or 2F) during the reproductive season (March-June). Birds received a photoperiod of 14L:10D and were fed ad libitum. Egg production and egg weight were recorded daily. Eggs were divided in 4 weight groups: EW1 =< 33 g, EW2 = 33-36 g, EW3 = 36-39 g and EW4 =≥ 39 g. Eggs were stored at 18 °C and classified in 3 egg storage groups: ES1 = 0-4, ES2 = 5-9 and ES3 = 10-15 days. Egg weight loss was recorded and distributed in 5 different classes: EWL1 =< 10%, EWL2 = 10-15%, EWL3 = 16-20%, EWL4 = 21-25%, EWL5 => 25%. Fertility, embryo mortality and hatchability were recorded. The mean values during the reproductive season were 82% fertility and 50% hatchability of fertile eggs. The best combination of fertility and hatchability values were recorded in EW2 and lower fertility was recorded in EW1 (P < 0.05). Hatchability decreased under 50% after 10 day storage period before incubation and the best hatchability was recorded in EWL1. The present results contribute to the knowledge on reproductive parameters necessary to improve the reproductive efficiency of this Italian breed within a conservation plan.

DNA fragmentation in chicken spermatozoa during cryopreservation.

Semen cryopreservation is fundamental both for the practice of artificial insemination, and for the conservation of genetic resources in cryobanks; nevertheless, there is still not an efficient standard freezing procedure assuring a steady and suitable level of fertility in fowl, and consequently there is no systematic use of frozen semen in the poultry industry. This study examined changes in motility (CASA), cell membrane integrity (Ethidium Bromide (EtBr) exclusion procedure and stress test) and DNA fragmentation (neutral comet assay) in fowl spermatozoa before, during and after cryopreservation and storage at -196 °C. An optimized comet assay for chicken semen was studied and applied to the analyses. Semen collected from 18 Mericanel della Brianza (local Italian breed) male chicken breeders was frozen in pellets and thawed in a water bath at 60 °C. Measurements were performed on fresh semen soon after dilution, after equilibration with 6% dimethylacetamide at 4 °C (processed semen) and after thawing. Sperm DNA damage occurred during cryopreservation of chicken semen and the proportion of spermatozoa with damaged DNA significantly increased from 6.2% in fresh and 6.4% in processed semen to 19.8% in frozen-thawed semen. The proportion of DNA in the comet tail of damaged spermatozoa was also significantly affected by cryopreservation, with an increase found from fresh (26.3%) to frozen-thawed (30.9%) sperm, whereas processed semen (30.1%) didn't show significant differences. The proportion of total membrane damaged spermatozoa (EtBr exclusion procedure) did not increase by 4 °C equilibration time, and greatly and significantly increased by cryopreservation; the values recorded in fresh, processed and frozen semen were 2.9, 5.6, and 66.7% respectively. As regards the proportion of damaged cells in the stress test, all values differed significantly (7.1% fresh semen, 11.7% processed semen, 63.7% frozen semen). Total motility was not affected by equilibration (52.1% fresh semen, 51.9% processed semen), whereas it decreased significantly after cryopreservation (19.8%). These results suggest a low sensitivity of frozen-thawed chicken spermatozoa to DNA fragmentation, therefore it should not be considered as a major cause of sperm injuries during cryopreservation.

Quality and lipid composition of spermatozoa in rabbits fed DHA and vitamin E rich diets.

The effects of fish oil (FO) and vitamin E (vE) dietary supplementation on semen quality, sperm susceptibility to lipid peroxidation, tocopherols content and fatty acid profiles were studied in rabbits. Fifty-two rabbit bucks randomly divided in four groups received a control diet and enriched diets containing either FO (1.5%, w/w), vE (200mg/kg) or both. Semen volume, concentration, motility and viability were analysed at various time-points and the lipid composition was assessed on sperm cells. The phospholipid fatty acid profile was determined: n-6 PUFA were the major fatty acids found, with a proportion of 42%, whereas the n-3 PUFA accounted for nearly 1%, mainly represented by C22:6n-3 (docosahexaenoic acid, DHA). FO supplementation produced a seven-fold increase in the content of DHA in sperm phospholipids and a comprehensive rearrangement of the phospholipid fatty acid composition, while an unexpected negative effect of feeding high level of vE on the proportion of total PUFA was found. Despite the remarkable changes observed in sperm lipid composition, semen quality parameters were not affected by the dietary treatments and the interaction between the two dietary supplements had a significant effect only on sperm concentration. An increase in semen production by ageing and a concomitant rise in sperm susceptibility to in vitro peroxidation was found. alpha- and delta-tocopherol, present in rabbit sperm in similar amount, were not affected by dietary treatment. delta-tocopherol content had a significant linear negative regression with age and showed a significant negative correlation with the susceptibility to peroxidation values.

Effect of docosahexaenoic acid and alpha-tocopherol enrichment in chicken sperm on semen quality, sperm lipid composition and susceptibility to peroxidation.

The aim of the present experiment was to study the effect of fish oil and Vitamin E rich diets on semen production, sperm functions and composition in broiler breeders. The following parameters were measured: semen volume and concentration, sperm motility and viability, sperm susceptibility to induced peroxidation, sperm lipid and alpha-tocopherol contents. Dietary n-3 PUFA were successfully transferred into spermatozoan phospholipid by fish oil feeding according to the following main features: (a) the C22:6n-3 and C22:5n - 3 contents were increased, but C22:4n-6 remained the peculiar and major polyunsaturate; (b) the content and proportion of total PUFA did not change; (c) the proportional increase of n-3 PUFA was compensated by the decrease of n-6 PUFA, an increase in the proportion of n-9 fatty acids was also found. The sperm content of alpha-tocopherol was doubled increasing the dietary availability of the vitamin to 300 mg/kg of feed. The specific n-3 PUFA and Vitamin E enrichment of chicken sperm affected cell functions. Significant interactions between the two treatments were also found for some parameters. The best sperm quality condition in control sperm (rich mainly in n-6 PUFA) was found supplying 200mg Vitamin E/kg of feed to the male breeders, and in contrast in n-3 rich sperm supplying 300 mg Vitamin E/kg.

Aquaporins are observed in the duct epithelia of the epididymal region of the large white turkey.

The cellular and molecular mechanisms regulating the reuptake of the testicular fluid supporting sperm exiting the testes in the bird are not known. The presence of aquaporins, proteins involved in transmembrane water transport, was investigated. Observations were limited to the ductuli efferentes, collecting ducts, and ductus epididymis. Interestingly all of these ducts were positive for aquaporins-2, -3, and -9 but not aquaporin-7. When positive, aquaporin was observed localized over the whole cell or the apical plasma membrane of the nonciliated cells and the apical plasma membrane and cilia of the ciliated cells. This study is the first to clearly demonstrate the presence of aquaporins-2, -3, and -9 in the epididymal region of any bird. We assume the aquaporins play a role in concentrating the sperm and in the promotion of sperm maturation in the epididymal region.

Localization of aquaporins in the sperm storage tubules in the turkey oviduct.

Oviductal sperm storage tubules (SST), located at the uterovaginal junction, are the primary site of sperm storage in turkeys. Sperm reside within these storage sites and may be released via a dynamic interaction between sperm mobility and a fluid current generated by the SST epithelial cells. In this study, aquaporins 2, 3, and 9 (proteins that form water channels in the plasmalemma of a variety of cells) were immunocytochemically localized within the apical aspect of the epithelial cells that form the SST. These observations support the contention that the SST epithelial cells are capable of water exchange that may interact with sperm residing within the SST.