E. Zanetti

Genetic variation and population structure of Italian native sheep breeds undergoing in situ conservation

The genetic variability and presence of population substructures in 4 native Northern Italian sheep breeds, Alpagota, Brogna, Foza, and Lamon, undergoing in situ conservation, and 1 widespread Italian breed, Bergamasca, were studied by investigating 19 microsatellite markers. The breeds showed considerable genetic variability in terms of number of alleles and heterozygosity, with the exception of Alpagota, which was the least variable (0.607). Nevertheless, a significant deficit of heterozygotes was observed in each breed due to rather increased levels of inbreeding or to the presence of population substructures, probably caused by increased genetic variation in the founder populations. The analyses evidenced clear genetic differentiation (F(ST) = 0.085), reduced levels of admixture, and presence of private alleles among the breeds, confirming their genetic uniqueness. In particular, according to Reynolds genetic distances, Alpagota was the most differentiated, perhaps because it had been bred mostly in a rather isolated area. Loss of any of the investigated breeds would result in a loss of genetic diversity ranging from 0.5 to 1.6% of the total observed gene diversity. Results supported the decision to safeguard these breeds as important reservoirs of genetic diversity and suggested breeding and mating practices to maintain variability and to overcome within-breed substructures.

Feasibility of the direct application of near-infrared reflectance spectroscopy on intact chicken breasts to predict meat color and physical traits

Physical and color characteristics of chicken meat were investigated on 193 animals by directly applying a fiberoptic probe to the breast muscle and using the visible-near-infrared (NIR) spectral range from 350 to 1,800 nm. Data on pH was recorded 48 h postmortem (pH); lightness (L*), redness (a*), and yellowness (b*) 48 h postmortem; thawing and cooking losses and shear force after freezing. Partial least squares regressions were performed using untreated data, raw absorbance data (log(1/R)), and multiplicative scatter correction plus first or second derivative spectra. Models were validated using full cross-validation, and their predictive ability was determined by root mean square error of cross-validation (RMSE(CV)) and correlation coefficient of cross-validation (r(cv)). Means (±SD) of pH, L*, a*, b*, thawing loss, cooking loss, and shear force were 5.83 ± 0.13, 44.54 ± 2.42, -1.90 ± 0.62, 3.21 ± 3.28, 4.84 ± 2.44%, 19.39 ± 2.95%, and 16.08 ± 3.83 N, respectively. The best prediction models were developed using log(1/R) spectra for b* (r(cv) = 0.93; RMSE(CV) = 1.16) and a* (r(cv) = 0.88; RMSE(CV) = 0.29), while a medium predictive ability of NIR was obtained for pH, L*, and thawing and cooking losses (r(cv) from 0.69 to 0.76; RMSE(CV) from 0.01 to 1.73). Finally, predicted model for shear force (r(cv) = 0.41; RMSE(CV) = 3.18) was unsatisfactory. Results suggest that NIR is a feasible technique for the assessment of several quality traits of intact breast muscle.

Variation of genetic diversity over time in local Italian chicken breeds undergoing in situ conservation

The aim of this study was to analyze the variation of different genetic diversity measures in 5 Italian local chicken breeds over a 4-yr period of conservation. The local breeds were Ermellinata di Rovigo, Padovana, Pépoi, Robusta Lionata, and Robusta Maculata. A total of 368 samples were chosen for the analysis among the birds hatched in the years 2002 and 2006. Genetic variation over 30 microsatellite loci was analyzed. All 30 microsatellites were polymorphic, with a total number of alleles equal to 251, a mean (±SD) of 8.367 ± 3.378 across populations and 3.233 ± 1.338 within population. After 4 yr of conservation, a loss of alleles occurred for all the microsatellites, with the exception of 4 loci. The total number of alleles and expected heterozygosity estimates significantly decreased during the 4 yr of conservation, whereas no significant differences were detected for the microsatellites polymorphism information content or for the observed heterozygosity estimates. A decrease of the inbreeding coefficient occurred for all the breeds, with the exception of Padovana and Robusta Lionata. All populations showed evidence of a persistent significant deviation from Hardy-Weinberg equilibrium caused by an excess of homozygotes, except Robusta Maculata, which reached the equilibrium in 2006. For Pépoi, Padovana, Ermellinata di Rovigo, and Robusta Maculata, molecular coancestry increased significantly (P < 0.001) after the 4 yr of conservation. No evident genetic structures were detectable within breed, both for 2002 and for 2006 individuals. However, a slight increase in the proportion of membership for each breed had occurred in the year 2006 compared with the data obtained in the year 2002. As consequence, within breed, individuals in 2006 appear more homogeneous, producing clearer, more distinctive and separated groups. Molecular markers analysis helped us monitor the genetic variability of local breeds involved in a conservation scheme, enabling the planning of new strategies for the improvement of in situ conservation schemes.

Using 2-DE for the differentiation of local chicken breeds

The aim of this study was to apply a proteomic approach for the characterisation of local chicken breeds. The experiment involved a total of 29 males of three local Italian chicken breeds: Pépoi, Padovana and Ermellinata di Rovigo. Sarcoplasmic protein fractions of breast muscle were analysed by 2‐DE. Image analysis followed by statistical analysis enabled to differentiate groups of individuals based on the similarities of protein expression. Individuals were distinguished into clusters and groups, corresponding to the breed of origin. Distances among individuals, calculated using data on spot volumes, were used to draw a neighbour‐joining tree, showing clear individual and breed grouping. The most relevant spots regarding breed differentiation were detected; 11 were identified by MS revealing preliminary evidences on the mechanisms of the breed differentiation process. The results evidenced the ability of proteomic analyses for the characterisation of chicken breeds.

Carcase characteristics and qualitative meat traits of three Italian local chicken breeds

1. An experiment involving 60 male chickens reared in an organic production system was carried out in order to investigate carcase characteristics and qualitative meat traits of three slow-growing Italian local breeds of chicken (Ermellinata, Padovana and Pépoi). 2. Chicks were randomly selected at hatch, raised together under the same conditions, slaughtered at 190 d of age, dissected for carcase traits, and meat (breast and thigh) stored for subsequent analysis of quality parameters. 3. Ermellinata (EA) chickens were significantly different from Padovana (PA) and Pépoi (PI) chickens for live, carcase and thigh weights. Breeds were also different for breast muscle protein content (EA > PI and PA), shear force (PA < EA and PI) and cooking loss (PI > PA and EA) values. 4. The CIE system values of lightness (L*), redness (a*), and yellowness (b*) evidenced a distinctive darker meat and lighter skin colour of PA breast meat. 5. Polyunsaturated fatty acids composition of breast meat was similar among the analysed breeds. EA had significantly higher saturated but significantly lower monounsaturated fatty acid contents than the other two breeds.

Survival analysis of piglet pre-weaning mortality

Abstract Survival analysis methodology was applied in order to analyse sources of variation of preweaning survival time and to estimate variance components using data from a crossbred piglets population. A frailty sire model was used with the litter effect treated as an additional random source of variation. All the variables considered had a significant effect on survivability: sex, cross-fostering, parity of the nurse-sow and litter size. The variance estimates of sire and litter were closed to 0.08 and 2 respectively and the heritability of pre-weaning survival was 0.03.

Estimation of genetic diversity over time in an in-situ marker assisted conservation scheme of local chicken breeds.

Abstract The aim of this research was to study the genetic variability of two local chicken breeds (Ermellinata di Rovigo and Robusta Maculata) involved in a conservation programme, over a period of three years. Samples were collected in 2002 and in 2005 and analysed using 15 microsatellite markers. Obtained results evidenced that Robusta Maculata was genetically less variable than Ermellinata di Rovigo even after three years of conservation activities. During the studied period the observed heterozigosity was maintained and the excess of homozygous animals found in both breeds in 2002 was strongly reduced in 2005. These findings suggested that the conservation activities helped to overcome the probable presence of population substructures and to limit inbreeding. Moreover, this study evidenced the usefulness of microsatellite markers to monitor genetic diversity in conservation programmes for animal breeds.

Genetic diversity in some local chicken breeds using microsatellite markers

Abstract Genetic relationships among Veneto native breeds of chickens were studied on the basis of microsatellites polymorphisms. A total of 100 DNA samples from 2 local chicken breeds (45 Robusta Lionata and 43 Robusta Maculata) and a commercial broiler line (12 Golden Comet) were analyzed using 19 microsatellite markers. The average number of alleles per locus was 4.05 and the expected heterozigosity resulted lower for the local breeds than the broiler line. The Robusta Lionata breed and the broiler line showed a significant deficit and excess of heterozygotes, respectively, deviating from Hardy-Weinberg equilibrium. Nei’s standard genetic distances corrected for bias due to sampling of individuals (Da), based on allele frequencies, were calculated among breeds. The local breeds resulted very similar confirming the same genetic origin. The results suggested that microsatellite markers are a useful tool for studying the genetic diversity among local chicken breeds.

A proteomic approach to study local chicken breeds characterization

Abstract Aim of this study is to apply a proteomic approach for characterization of local chicken breeds. The experiment involved a total of 29 males of Pépoi, Padovana, and Ermellinata local chicken breeds. Sarcoplasmic protein fractions of breast muscle were analysed by bidimensional electrophoresis. Image analysis followed by statistical analysis enabled to differentiate groups of individuals on the similarities of protein expression. Individuals were distinguished into clusters and groups, corresponding to the breed of origin. Results evidenced a possible utilisation of proteomic approach in the field of breed characterization studies as an alternative to genomic analyses performed using molecular markers, both for breed and product traceability purposes.

Development and validation of a novel SNP panel for the genetic characterization of Italian chicken breeds by next-generation sequencing discovery and array genotyping

ABSTRACT The aim of this study was to compare the intra and inter genetic variability and population structure of 7 indigenous chicken breeds of the Veneto region, through a novel panel of 64 SNP, each located in an exonic region and mostly on different chromosomes. A total of 753 blood samples from 7 local chicken breeds (Ermellinata di Rovigo, Millefiori di Lonigo, Polverara, Pepòi, Robusta Lionata, Robusta Maculata, and Padovana) was collected and analyzed. Two strains of Polverara (Nera and Bianca) and Padovana (Dorata and Camosciata) were included in the study. The observed heterozygosity ranged from 0.124 (Pèpoi) to 0.244 (Ermellinata di Rovigo), and the expected heterozygosity varied from 0.132 (Millefiori di Lonigo) to 0.300 (Ermellinata di Rovigo). Global FIS results (0.114) indicated a low‐medium inbreeding effect, with values ranging from 0.008 (Millefiori di Lonigo) to 0.223 (Ermellinata di Rovigo). Pairwise FST values (0.167) for all populations ranged from 0.020 (Polverara Nera and Polverara Bianca) to 0.193 (Robusta Lionata and Polverara Nera), indicating that the studied breeds were genetically highly differentiated. The software STRUCTURE was used to detect the presence of population substructures, and the most probable number of clusters (K) of the 10 chicken populations was at K = 8. The affiliation was successful in all Veneto chicken breeds. The present SNP marker results, compared with previous data obtained using microsatellites, provided a reliable estimate of genetic diversity within and between the studied breeds, and demonstrated the utility of the proposed panel as a rapid, efficient, and cost‐effective tool for periodical monitoring of the genetic variability among poultry populations. In addition, the present SNP panel could represent a resource for a systematic approach with relevant impact on breeding program decisions and could turn out to be a reliable tool for genetic traceability of indigenous chicken meat. Adoption of a periodical monitoring system of genetic diversity is a fundamental tool in conservation actions and should increase the value of typical and niche products.