Emanuela Sgreccia

Luca Pasquini

Gianfranco Ercolani

Philippe Knauth

Maria Luisa

Di Vona
email: divona@uniroma2.it

Maria Luisa Di Vona

Stimuli-responsive amphoteric ion exchange polymers bearing carboxylic and amine groups grafted to a cross-linkable silica network

Keywords: ionomers, polysulfone, pH-dependence, sol-gel, amphoteric

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Polyelectrolytes are a fascinating class of macromolecules with applications in different fields comprising biomedical and soft matter sciences. [START_REF] Ulrich | Formation of Complexes between Nanoparticles and Weak Polyampholyte Chains. Monte Carlo Simulations[END_REF][START_REF] Schachschal | Polyampholyte Microgels with Anionic Core and Cationic Shell[END_REF] Electrostatic interactions between charged groups and/or solvents lead to a considerably different behaviour compared to uncharged polymers given the dominant role played by Coulomb interactions. [START_REF] Ciferri | Natural and synthetic polyampholytes, 1(a) theory and basic structures[END_REF] Amphoteric ionomers are a particular class of polyelectrolytes containing both acidic and basic groups in their structure. [START_REF] Xu | Ion exchange membranes: State of their development and perspective[END_REF] Typically, amphoteric polymers bear fixed positive and negative charges formed by weak basic and acidic moieties.

Among synthetic polyelectrolytes, amphoteric polymers are close to biological macromolecules.

They may provide useful analogues of proteins and are important to model some properties and functions of biopolymers, such as protein folding and enzymatic activity. They are also relevant in separation technologies [START_REF] Lin | Extraction of Amphoteric Amino Acid by Bipolar Membrane Electrodialysis: Methionine Acid as a Case Study[END_REF][START_REF] Miao | Amphoteric composite membranes for nanofiltration prepared from sulfated chitosan crosslinked with hexamethylene diisocyanate[END_REF][START_REF] Nesterenko | Zwitterionic ion-exchangers in ion chromatography: A review of recent developments[END_REF][START_REF] Cheng | High-performance protein separation by ion exchange membrane partitioned free-flow isoelectric focusing system[END_REF] and drug delivery, [START_REF] Ortiz-Collazos | Interaction of levofloxacin with lung surfactant at the air-water interface[END_REF][START_REF] Atamanyuk | Vectorization Efforts To Increase Gram-Negative Intracellular Drug Concentration: A Case Study on HldE-K Inhibitors[END_REF][START_REF] Ali | A pH-Tunable Nanofluidic Diode with a Broad Range of Rectifying Properties[END_REF] but also in electrochemical energy technologies, such as polymer electrolyte membrane fuel cells [START_REF] Tadavani | Synergistic Behavior of Phosphonated and Sulfonated Groups on Proton Conductivity and Their Performance for High-Temperature Proton Exchange Membrane Fuel Cells (PEMFCs)[END_REF][START_REF] Knauth | Comparative study of the cation permeability of protonic, anionic and ampholytic membranes[END_REF] and redox flow batteries. [START_REF] Liao | Fluoro-methyl sulfonated poly(arylene ether ketone-co-benzimidazole) amphoteric ion-exchange membranes for vanadium redox flow battery[END_REF][START_REF] Lee | A novel amphoteric ion-exchange membrane prepared by the pore-filling technique for vanadium redox flow batteries[END_REF][START_REF] Liao | Ultra-low vanadium ion diffusion amphoteric ionexchange membranes for all-vanadium redox flow batteries[END_REF] It was also reported that polymers with weak acid or basic groups can activate water splitting in bipolar membranes, considering that water splitting strongly depends on the interface characteristics. [START_REF] Kang | Enhancement of water splitting in bipolar membranes by optimized composite anion-exchange layer and alkali-treated polyacrylonitrile catalytic junction[END_REF][START_REF] Balster | Tailoring the interface layer of the bipolar membrane[END_REF][START_REF] Zabolotsky | Modification of asymmetric bipolar membranes by functionalized hyperbranched polymers and their investigation during pH correction of diluted electrolytes solutions by electrodialysis[END_REF][START_REF] Simons | Strong electric-field effects on proton-transfer between membrane-bound amines and water[END_REF][START_REF] Greben | Influence of ionite nature on physicochemical properties of bipolar ion-exchange membranes[END_REF] Different polyelectrolytes were used for this aim: anion exchange membranes with secondary amine or tertiary ammonium groups or proton exchange polymers with weak acidic groups. [START_REF] Zabolotsky | Modification of asymmetric bipolar membranes by functionalized hyperbranched polymers and their investigation during pH correction of diluted electrolytes solutions by electrodialysis[END_REF] A peculiarity of amphoteric polymers is that the electrostatic interaction can be varied changing pH and ion concentrations modifying the kind and nature of the interfaces with surrounding phases. For example, their adsorption on charged surfaces was studied in depth in the last years. [START_REF] Kudaibergenov | Natural and synthetic polyampholytes, 2(a) functions and applications[END_REF][START_REF] Kudaibergenov | Preparation and characterization of novel polymeric betaines based on aminocrotonates[END_REF] Synthetic ampholytic ionomers are often prepared by polymerization of alkenyl moieties [START_REF] Pu | Synthesis and properties of amphoteric copolymer of 5vinyltetrazole and vinylbenzyl phosphonic acid[END_REF][START_REF] Bozkurt | Copolymers of 4(5)-vinylimidazole and ethyleneglycol methacrylate phosphate: Synthesis and proton conductivity properties[END_REF] or by blending acidic and basic polymers. [START_REF] Karadedeli | Proton conduction in adipic acid/benzimidazole hybrid electrolytes[END_REF] In the past, we synthetized amphoteric aromatic polymers starting by a chlorosulfonated synthon. [START_REF] Narducci | Low-Permeability Poly(ether Ether Ketone)-Based Ampholytic Membranes[END_REF] In this paper, we report the synthesis and the characterization of a novel class of amphoteric ionomers where the acidic and basic groups are part of an inorganic network forming ionic clusters grafted on a polymeric matrix.

These class II hybrids present covalent bonds among the organic and inorganic fragments. [START_REF] Sanchez | Design of functional nano-structured materials through the use of controlled hybrid organic-inorganic interfaces[END_REF] Hybrid polymers represent a promising class of materials, which allow regulating a suitable ratio between hydrophilic and hydrophobic domains; the resulting ionomers intrinsically retain chemical and physical properties of both inorganic and organic components. Generally, expected improvements of ionomer properties by inorganic groups include an increased thermal and mechanical stability, the achievement of an interconnected porous network, or specific chemical properties.

We have prepared different class II hybrids, following several strategies. [START_REF] Licoccia | SPPSU-based hybrid proton conducting polymeric electrolytes for intermediate temperature PEMFCs[END_REF][START_REF] Vona | Organic-inorganic hybrid membranes based on sulfonated polyaryl-ether-ketones: Correlation between water uptake and electrical conductivity[END_REF][START_REF] Sgreccia | Mechanical properties of hybrid proton conducting polymer blends based on sulfonated polyetheretherketones[END_REF][START_REF] Sgreccia | Selfassembled nanocomposite organic-inorganic proton conducting sulfonated poly-ether-ether-ketone (SPEEK)-based membranes: Optimized mechanical, thermal and electrical properties[END_REF][START_REF] Sgreccia | Hybrid composite membranes based on SPEEK and functionalized PPSU for PEM fuel cells[END_REF][START_REF] Knauth | Water Activity Coefficient and Proton Mobility in Hydrated Acidic Polymers[END_REF] The sol-gel process represents a versatile synthetic approach to tailor structures at a molecular level controlling the amount of different components with fine tuning of the inorganic clusters. The presence of functionalizable Si-OH groups in the hybrid ionomers synthesized in this work opens flexible and simple routes to tailor reticulation and mesoporosity, or to bond molecules modulating their chemical and biological properties.

All components of the hybrid ionomers synthesized in this work, polysulfone (PSU), amine and carboxylic acid groups and silanol moieties are biocompatible and can be employed in the biological field. The acid-base interaction in the amphoteric polymers is very important for the ion transport and we paid particular attention to the stimuli-sensitive properties as a function of pH.

An understanding of the physico-chemical properties of such polyampholytes can be relevant for many applications.

Experimental

Materials

Polysulfone (MW = 55500 g/mol, PSU) was obtained from Solvay. 3-(2-aminoethylamino) propyldimethoxymethylsilane (AEAPS), 3-cyanopropyltrichlorosilane (CNSi) and all other chemicals were reagent grade and were used as received from Aldrich.

Synthesis of hybrid carboxylated and aminated PSU (PSU-SiCA)

-Aminated PSU (PSU-CH 2 AEAPS)

The chloromethylation of PSU was reported elsewhere. [START_REF] Vona | Anion-conducting ionomers: Study of type of functionalizing amine and macromolecular cross-linking[END_REF][START_REF] Pasquini | Effects of anion substitution on hydration, ionic conductivity and mechanical properties of anion-exchange membranes[END_REF][START_REF] Vona | Anionic conducting composite membranes based on aromatic polymer and layered double hydroxides[END_REF] PSU-CH 2 Cl (degree of chloromethylation = 0.7; 1.08 meq) was dissolved in 50 mL of anhydrous THF at RT under N 2 .

Anhydrous sodium acetate and 20 mL of a solution of AEAPS in THF (eq PSU-CH2Cl :eq AcONa : eq AEAPS = 1:1:1) were added and the solution heated at reflux under stirring for 48h. After this time, a sample was taken and directly dried for NMR and FTIR analyses (Supplementary Materials). -Carboxylated and aminated PSU (PSU-CH 2 AEAPS-CNSi) Glacial AcOH was added to the solution containing PSU-CH 2 AEAPS and the reaction was kept at reflux for other 4h under N 2 . The solution was cooled to RT and CNSi (eq PSU-CH2Cl :eq AcOH :eq CNSi = 1:3:1) was directly added. The reaction was kept at reflux and left for 48h under stirring. A second batch was taken, dried, and analysed by FTIR (Supplementary Materials).

The muddy dark orange solution was precipitated in ethanol and washed several times with water.

PSU-CH 2 AEAPS-CNSi was hydrolysed with 0.1 M NaOH during 4h at reflux. After this time, the powder was washed with water and dried at 60°C for 3 days.

The reaction was repeated using PSU-CH 2 Cl DCM = 0.5 and two moles of CNSi to balance the acid and base groups (eq PSU-CH2Cl :eq AcONa :eq AEAPS: eq AcOH :eq CNSi = 1:1:1:3:2). The two organic-inorganic amphoteric ionomers are in the following called PSU-SiCA(1:2) and PSU-SiCA(2:2), indicating the ratio between acid and basic groups.

Titration

The acid form of the two amphoteric ionomers was obtained by immersion under stirring in 2 M HCl during 24 h. Powders were filtered, repeatedly washed with water and then dried in the oven at 60 °C under vacuum and one week over P 2 O 5 . A weighed quantity of the dry samples was immersed in water and potentiometrically titrated with 0.022 M NaOH under nitrogen. The pH values were measured after equilibration of the mixture; this process can take up to 16 h. The titration gives the acid and base constants and the isoelectric point, as discussed in the results section.

The total ion exchange capacity (IEC) was determined by back titration. The basic form was obtained by immersion under stirring in NaOH 2M during 24 h. After washing with water, the samples were placed in 5 mL of 0.1 M HCl for 24 h, and then the solution was back titrated with NaOH 0.02 M. The results are reported in Table 1.

Hydration measurements

The water uptake at 25 °C was determined by exposition of powders to saturated water vapour during 1 week. The sample mass (m wet ) was determined by rapid weighing in a closed vessel. The dry sample mass was measured after drying over P 2 O 5 for 24 h (m dry ).

The water uptake WU was calculated from the equation:

(1)

The hydration isotherm was determined at 25 °C between 5 and 95% relative humidity (RH) using a thermogravimetric analyzer TA Q5000. The sample mass was about 7 mg; the sample was dried in situ 3 hours at 80 °C before the measurements. RH was then changed by 10 % increments during 3 hours. The hysteresis was studied by measurements in the opposite direction, between 95 and 5% RH.

Conductivity measurements

The ionic conductivity was determined by electrochemical impedance spectroscopy EIS (Parstat 4000) in fully humidified conditions using a Swagelok cell with cylindrical stainless steel electrodes of 6 mm diameter. The ac voltage amplitude was 20 mV and the frequency range was between 10 Hz and 6 MHz.

Sample powders were compressed to dense pellets under 1 MPa pressure at room temperature.

The samples were cut in the middle part of the pellets in order to avoid thickness changes at the periphery. The through-plane resistance of the samples R mat was determined from a non-linear least-square fitting of Nyquist plots using the Zfit program. The ion conductivity σ was calculated using the following equation:

A R d mat    ( 2
)
where d is the sample thickness and A = 0.28 cm 2 the electrode area.

Scanning Electron Microscopy (SEM)

The scanning electron micrographs were recorded on a Hitachi S-2460N microscope, using an acceleration voltage of 20 kV and an emission current of 90 µA. The reaction routes are reported in Figure 1a. The synthesis was made in anhydrous THF, to avoid the sol-gel self-condensation of AEAPS. The first step of the amphoteric-PSU synthesis is the formation of the aminated derivative. The reaction is a nucleophilic bimolecular substitution (S N 2)

Results and Discussion

Synthetic strategy and structural characterization

where the N atom acts as nucleophile. Two different reaction centres are present in AEAPS: primary and secondary amine. The nucleophilic character of nitrogen depends on many factors, among them basicity, partial electrical charge, solvation, steric phenomena and polarizability. The parameter defined by Swain and Scott for nucleophilic attack upon saturated carbon atoms shows that the amine reactivity lies in the order tertiary > secondary > primary justifying the formation of the tertiary amine (Figure 1). [START_REF] Bunting | Nucleophilicity towards a saturated carbon-atom -rate constants for the aminolysis of methyl 4-nitrobenzenesulfonate in aqueous-solution -a comparison of the n and n-+ parameters for amine nucleophilicity[END_REF][START_REF] Swain | Quantitative Correlation of Relative Rates. Comparison of Hydroxide Ion with Other Nucleophilic Reagents toward Alkyl Halides, Esters, Epoxides and Acyl Halides1[END_REF] Acetic acid or acetate, added during the first step, can have an additional effect: some AcO groups can weakly bond to Si alkoxide via a fivefold coordination with silicon [START_REF] Robert | Stereochemistry at Silicon[END_REF] forming an alcohol (Eq. 3). This equilibrium is shifted towards the left and only few acetate groups are consumed, although heating the solution under reflux moves the equilibrium towards the right, because the reaction is endothermic. [START_REF] Sanchez | Chemical modification of alkoxide precursors[END_REF] Si(OR

) n + x AcOH  Si(OR) n-x (OAc) x + x ROH (3)
The second step is a non-hydrolytic reaction between silicon alkoxides and silicon chlorides that is generally carried out around 100 °C or more and involves the elimination of alkyl halide. [START_REF] Mutin | Nonhydrolytic Processing of Oxide-Based Materials: Simple Routes to Control Homogeneity, Morphology, and Nanostructure[END_REF] In the case of metal acetates, the reaction is supposed to occur also at a lower temperature (Eq. 4). In our case, to favour the reaction in THF and to protect amino groups, acetic acid was added before the reaction with CNSi to form Si-OAc.

Si-Cl + Si-OAc  Si-O-Si + AcCl ( 4
)
The reaction is a nucleophilic substitution involving SiO ( -Ac ( polarization leading to an acyl cation (CH 3 CO + ). The last step of the synthesis of PSU-SiCA is the hydrolysis in basic condition of the cyano groups (Figure 1a). This step leads also to the formation of some cross-links between the repeat units creating a silica network (Figure 1c).

Two different molar ratios AEAPS:CNSi were used 1:1 and 1:2. In the first case, there is an excess of amine groups, in the second case, the quantity NH/CO 2 H is equal.

Based on the structures of PSU-SiCA(1:2) and PSU-SiCA(2:2) in Figure 1, the compositional characteristics of the samples are reported in Table 1. The degrees of functionalization were assessed from elemental analysis and thermogravimetry (TGA, see below for discussion). The molecular weights were calculated according to Figure 1 for a degree of functionalization of 0.5; however, the partial cross-linking of the silica network could not be taken into account in this simple calculation. The calculated total IEC is compared with data from acid-base titration. The total ion exchange capacity of the two samples (Table 1) was obtained by back-titration with NaOH of the acidic form (made according to eq. 5). The obtained IEC are consistent with a degree of functionalization 0.5.

Sample

R-NH-

….-COO --+ 2 H +  R-NH 2 + -….-COOH (5)
The insolubility of the two samples in organic solvents, also in acid form, containing 6.1 % PSU-SiCA(2:2) and 4.5 % PSU-SiCA(1:2) of Si respectively, [START_REF] Weldes | PROPERTIES OF SOLUBLE SILICATES[END_REF] can be attributed to the presence of crosslinks between the silanol groups. The low IEC value measured by titration for the sample PSU-SiCA(2:2) (Table 1) can be related to this formation of a cross-linked polymeric network, evidently facilitated by the presence of another silanol group.

Thermogravimetric analysis

The thermogravimetric curve of PSU-SiCA(1:2) (Figure 2) shows two main mass loss peaks. The first one starting above 330 °C can be attributed to the loss of amine groups and the complete side chain. This is in accordance with the literature, reporting that aminated PSU loses the amine group around 330 °C. [START_REF] Jyothi | Aminated polysulfone/TiO2 composite membranes for an effective removal of Cr(VI)[END_REF] The calculated mass loss corresponding to the loss of the branched side chain (MW = 338 g/mol, degree of functionalization = 0.5) is around 27 %, in good agreement with the experiment. A remarkable effect of the inorganic component is the stabilization of the carboxylic groups, because in carboxylated PSU the degradation of CO 2 H starts already around 290°C, [START_REF] Guiver | Synthesis and Characterization of Carboxylated Polysulfones[END_REF] The decomposition of the PSU main chain (MW = 442 g/mol) above 450 °C is in line with previous experiments. Finally, the residue above this temperature, which is formed by SiO 2 , should give a residual mass percentage of 10% (M(SiO 2 )/M(repeat unit)) in excellent agreement with the observation. Altogether, the thermogravimetric analysis confirms the composition of the hybrid sample. [START_REF] Knauth | Thermogravimetric analysis of SPEEK membranes: Thermal stability, degree of sulfonation and cross-linking reaction[END_REF] Figure 2. Thermogravimetric curve of the decomposition of PSU-SiCA(1:2). T / °C Ampholytic ionomers show many properties, such as ionic conductivity and swelling behaviour, which are strongly pH-dependent, because they are sensitive to the ionization of functional groups in the membrane matrix. In the isoelectric region, the membrane deswells in length and mass and the electric resistance increases. The ion selectivity is determined by the sign of fixed charges. If the charge is positive, the membrane is anion-selective; if the charge is negative, it is cationselective. Therefore, the membrane is anion selective in acidic conditions, whereas it is cation selective in basic conditions. If both acidic and basic functional groups are ionized, the average distance between opposite charges is important.

Determination of the acid and base constants by titration

Figure 3 shows schematically the different forms of amphoteric polymers as a function of pH. At low pH, the amino moieties have a positive charge while the carboxylic groups are essentially undissociated. Increasing the pH, the carboxylic acids start to dissociate due to their higher acid constants with respect to ammonium moieties (typically 10 -5 vs 10 -9 ). When all carboxylic groups have lost their protons, the ammonium groups dissociate with further pH increase.

+ + + + - - - - 2
The following equation relates the isoelectric point (pI) to the acid constants when the number of acid (carboxylic) and basic (amine) groups is equal: [START_REF] Ciferri | Natural and synthetic polyampholytes, 1(a) theory and basic structures[END_REF] Here pK a1 is the acid constant of the carboxylic groups, whereas pK a2 is the acid constant of the ammonium groups.

From the potentiometric titration curve for PSU-SiCA(2:2) (Supplementary Materials) a pI = 5.7 can be estimated from the inflection point, which corresponds to the neutralization of the two carboxylic groups. The acid constant of the second carboxylic group can be calculated from the pH at half neutralization. From the titration curve, one can read pK a1 = pH = 4.4, which is realistic for a carboxylic acid. Using equation 6, one gets for the acid constant of the first ammonium group: pK a2 = 7.0, which corresponds to a basic constant pK b2 = 7.0 for the secondary amine group (conjugated base). This pK a2 value can be confronted with the value of the curve at half neutralization of the first ammonium group giving pK a2 = 7.2 in good agreement with the above value. Finally, the pH value at half neutralization gives the acid constant of the second ammonium group: pK a3 = 7.7

corresponding to a basic constant pK b3 = 6.3 of the conjugated primary amine. Figure 4 shows the swelling-deswelling behaviour of ampholytic ionomers in different pH conditions. Polyampholyte chains assume a globular conformation and reduced solubility at pH = pI, which has been attributed to an intramolecular charge screening effect. [START_REF] Ciferri | Natural and synthetic polyampholytes, 1(a) theory and basic structures[END_REF][START_REF] Patrickios | POLYPEPTIDE AMINO-ACID-COMPOSITION AND ISOELECTRIC POINT .1. A CLOSED-FORM APPROXIMATION[END_REF] A minimum swelling value is observed at the isoelectric point. The swelling increases when the concentration of the grafted anions and cations is unbalanced.

Water Uptake

The water uptake measured at 25 °C for the two compounds in acidic, basic and zwitterionic forms (Figure 5) shows the lowest values for the zwitterionic form in agreement with the expectation.

The generally low water uptake is probably related to the low dissociation of carboxylic acids, so that only few hydrophilic ionized groups are present in the macromolecule. The acid forms, containing mobile chloride anions and grafted ammonium and carboxylic acid groups, present a higher water uptake than the basic forms, containing mobile sodium cations and grafted carboxylate and amine groups. The higher hydration observed for PSU-SiCA(1:2) is consistent with its higher IEC (Table 1).

The hydration isotherms of the two acidic forms are shown in Figure 6. The water uptake is relatively low, below 10%, consistent with the data reported in Figure 5. The higher water uptake observed for PSU-SiCA(1:2) is in agreement with the data in saturated water vapour. The aspect of the hydration isotherms is in line with previous experiments on proton-conducting ionomers [START_REF] Knauth | Chemomechanics of acidic ionomers: Hydration isotherms and physical model[END_REF] and corresponds to type 2 BET adsorption. [START_REF] Thommes | Physisorption of gases, with special reference to the evaluation of surface area and pore size distribution (IUPAC Technical Report)[END_REF] Whereas the initial part of the isotherms (up to about 70 %RH) can be attributed to formation of a monolayer of water molecules, the final part corresponds to multilayers of water in pores of the ionomers. The hysteresis of the desorption part is classical for a material with a large quantity of mesopores, due to the presence of the inorganic network. This opportunity can be further exploited for the synthesis of materials with controlled mesoporosity.

Ionic conductivity

At low and high pH (Figure 4), polyampholytes can be considered as ionomers with mobile anions or cations, respectively. The behaviour at the pI depends on the distance between the grafted charged groups. If the opposite ions are near enough (typically below the Bjerrum radius, i.e. around 0.7 nm) they neutralize each other (or form ion pairs) by Coulombic attraction and the ionic conductivity should present a minimum value. If the distance between charges is high, anions and cations might both contribute to the conductivity. [START_REF] Kudaibergenov | Amphoteric nano-, micro-, and macrogels, membranes, and thin films[END_REF][START_REF] Korosy | AMPHOTERIC ION-PERMSELECTIVE MEMBRANE[END_REF] The ionic conductivity of PSU-SiCA(1:2) and PSU-SiCA(2:2) samples in acidic, basic and zwitterionic form at temperatures between 298 and 353 K is reported in Table 2. The ionic conductivity of PSU-SiCA(1:2) is consistently higher than that of PSU-SiCA(2:2), due to a higher IEC of the former. In agreement with the expectation, the zwitterionic forms show the minimum of ionic conductivity; the highest ionic conductivity is observed for the acidic forms, where chloride ions conduct together with few protons from partially dissociated carboxylic acid groups. The variations of ionic conductivity are accompanied by differences in activation energies.

The activation energy calculated from the data for the two acidic forms is (0.25 ± 0.02) eV, consistent with a predominant migration of chloride ions. Similar values were reported for other anion-conducting ionomers [START_REF] Braglia | Electrochemical synthesis of thin, dense, and conformal anion exchange membranes with quaternary ammonium groups[END_REF][START_REF] Vlaev | Concentration dependence of the activation energy of conductivity in aqueous sodium selenite and potassium tellurite[END_REF] . In the basic forms, sodium ions are the predominant ionic carriers and the activation energy calculated for the two basic forms is higher (0.37 ± 0.05) eV.

Slightly lower values were reported for sodium ion transport in aqueous solution or in Nafion. [START_REF] Vlaev | Concentration dependence of the activation energy of conductivity in aqueous sodium selenite and potassium tellurite[END_REF][START_REF] Hongsirikarn | Effect of cations (Na+, Ca2+, Fe3+) on the conductivity of a Nafion membrane[END_REF] Finally, the activation energy is highest for the zwitterionic form: (0.44 ± 0.03) eV.

More than the absolute values of ionic conductivity, which remain modest, the versatility and the pH-stimulus dependence are of real interest.

To study the stability of these ampholytic polymers in alkaline solution, the PSU-SiCA(2:2) powder was treated with 2M NaOH at 80 °C during one week. After this harsh treatment, the FTIR spectrum did not show appreciable differences. The IEC of treated samples was also measured by We tested the opportunity of preferred deposition on oxide substrates with basic or acidic properties; the interfacial variability is one of the most attractive features of amphoteric polymers.

Figure 7 shows SEM images of PSU-SiCA(2:2) deposited on SiO 2 and Al 2 O 3 substrates . The appearance is typical of hybrid materials with an inorganic (silica) part. The basic compound is deposited only on the acidic substrate (SiO 2 ), while the acidic form is observed only on the basic substrate (Al 2 O 3 ). This observation is consistent with the excess charge of the ionomers. The basic form presents an excess of grafted negative charges, due to the carboxylate anions, which are attracted by the surface protons of the acidic silica substrate. The acidic form contains an excess of positive charges due to the anchored ammonium cations, which are interacting with the hydroxide surface groups of the basic alumina substrate. The zwitterionic form does not deposit on both substrates, because it does not present excess charges and attractive and repulsive interactions are balanced (Figure 7a). These experiments show impressively how the interface potential modifies the adsorption and the phase deposition of ampholytic polymers, which allows a controlled synthesis of complex material/material architectures.

Conclusions

We have synthesized hybrid organic-inorganic amphoteric ionomers by non-hydrolytic sol-gel chemistry. They include a silica-based inorganic side chain with grafted functional groups, which allows developing materials with controlled microporosity and pH-sensitivity. Thermogravimetric and elemental analysis confirmed the presence of the inorganic component and helped to assign the degree of functionalization. The isoelectric point and the various acid and base constants were determined by acid-base titration. The deposition on substrates with basic and acidic behaviour shows the influence of the interfacial potential on adsorption and phase deposition allowing the controlled design of complex architectures. The pH-dependent variations of various properties, such as hydration and ionic conductivity of acidic, basic and zwitterionic forms, gives the opportunity to tune and optimize these properties by an external stimulus.

Figure 1

 1 Figure 1. a) Reaction pathways for the synthesis of PSU-SiCA(2:2); b) Repeat unit of PSU-SiCA(1:2); c) Schematic of cross-linking between silica networks.

Figure 3 .

 3 Figure 3. Ion conduction in amphoteric ionomers as a function of pH (adjusted by HCl and NaOH). The zwitterionic form is observed at an intermediate pH.

Figure 4 .

 4 Figure 4. Schematic representation of the swelling-deswelling behaviour of PSU-SiCA(2:2) as a function of pH. Color code: Si: green, O: red, N: blue, S: yellow, C: grey.

Figure 5 .

 5 Figure 5. Water uptake in saturated humidity (RH = 100 %) at 25 °C of PSU-SiCA(1:2) and PSU-SiCA(2:2) in acidic (red), basic (blue) and zwitterionic (grey) forms.

Figure 6 .

 6 Figure 6. Hydration isotherms of PSU-SiCA(1:2) (squares and dotted lines) and PSU-SiCA(2:2) (dots and full lines). The red curves correspond to water desorption.

Figure 7

 7 Figure 7. a) Schematic interface interaction; SEM micrographs of PSU-SiCA(2:2) deposited on b) SiO 2 (basic form) and c) Al 2 O 3 (acid form).

Table 1 .

 1 Degree of functionalization, molecular weight (MW) and ion exchange capacity (IEC) of hybrid amphoteric polymers.

		Degree of	Calculated MW	Calculated	Total IEC
		Functionalization	repeat unit	total IEC	(titration)
			g/mol	meq/g	meq/g
	PSU-SiCA(1:2)	0.5 (TGA)	612	1.64	1.80
	PSU-SiCA(2:2)	0.5 (el. analysis)	686	1.45	1.20

The elemental analysis (mass%) performed on PSU-SiCA(2:2) gave results in good agreement with a degree of functionalization 0.5, corresponding to a formula C 34.5 H 39.5 NO 9 SSi 1.5 : C calculated 60.3, measured 62.1, N calculated 2.0, measured 2.2, Si calculated 6.1 measured 6.5.

Table 2

 2

	. Ionic conductivity of PSU-SiCA(1:2) and PSU-SiCA(2:2) at temperatures between 298 and
	353 K in acidic, basic and zwitterionic forms.
	Form	T / K	PSU-SiCA(1:2) S/cm	PSU-SiCA(2:2) S/cm
	Acidic	298	2.6 10 -5	1.5 10 -5
		313	4.4 10 -5	2.9 10 -5
		333	-	3.7 10 -5
		353	-	7.1 10 -5
	Basic	298	7.8 10 -6	2.0 10 -6
		313	1.6 10 -5	4.4 10 -6
		333	3.3 10 -5	-
		353	5.4 10 -5	-
	Zwitterionic	298	5.5 10 -7	4.5 10 -7
		313	1.5 10 -6	4.7 10 -7
		333	5.5 10 -6	6.1 10 -7
		353	8.8 10 -6	6.5 10 -6

μm a)

Acknowledgments

The authors thank Dr. Roberto Volterri for his valuable technical assistance for the SEM experiments.

Funding sources

This work was supported by the European Union in the framework of the Horizon 2020 project "CREATE" (GA 721065).

Data availability

The raw/processed data required to reproduce these findings cannot be shared at this time as the data also forms part of an ongoing study.


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Keywords: carcass, frozen semen, genetic trend, growth, swine

niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

INTRODUCTION

The major pig populations have been selected for decades to improve growth efficiency, carcass quality, and, over the last 15 yr, prolificacy in maternal lines [START_REF] Ollivier | Genetic Improvement of the Pig[END_REF][START_REF] Tribout | Continuous genetic evaluation of on farm and station tested pigs for production and reproduction traits in France[END_REF]. Large improvements have generally been obtained for the main selected traits (e.g., [START_REF] Ducos | Utilisation d'un modèle animal multi-caractères pour estimer l'évolution génétique de six caractères de croissance, carcasse et qualité de la viande entre 1977 et 1990, dans les races porcines Large White et Landrace français[END_REF][START_REF] Chen | Genetic parameters and trends for lean growth rate and its components in U.S. Yorkshire, Duroc, Hampshire, and Landrace pigs[END_REF]. Genetic trends for these traits, in particular selection criteria that are routinely measured, can easily be estimated using BLUP genetic evaluation systems, provided that all the information pertaining to the selection process is included in the analyses [START_REF] Im | Likelihood inferences in animal breeding under selection: A missing-data theory view point[END_REF]. However, selection is likely to affect a much larger number of traits that are genetically correlated with those used as selection criteria, but genetic trends for these traits cannot be estimated because of a lack of data. As suggested by [START_REF] Smith | Use of stored frozen semen and embryos to measure genetic trends in farm livestock[END_REF], this difficulty can be circumvented by storing gametes or embryos and using them after some time to produce groups of pigs that can be compared with more recent populations and that can provide estimates of genetic trends for a large number of traits.

A stock of frozen semen of Large White (LW) boars born in 1977 was collected for this purpose at the end of the 1970s. A part of the stock was used to estimate genetic trends for performance and meat quality traits after 5 and 10 yr of selection [START_REF] Molénat | Utilisation de la semence congelée pour l'estimation du progrès génétique sur les caractères de production chez le porc[END_REF][START_REF] Ollivier | Progrès génétique des porcs Large White et Landrace français de 1977 à 1987[END_REF]. Twelve years later, it was decided to repeat the experiment in the LW breed so as to estimate genetics trends after 21 yr of selection for a wider range of production, lean and fat tissue quality, and reproduction traits. This paper presents the experimental design and the results for growth and carcass traits.

MATERIALS AND METHODS

Animals used in this study were raised and slaughtered in accredited slaughterhouses according to the protection of animals rules defined in the French law (Code Rural, articles R214-64 to R214-71; http://www. legifrance.gouv.fr).

Brief History of Selection in the French LW Breed from 1977 to 1998

As with most commercial pig populations, the breeding goal of the French LW population has evolved several times in recent decades. Until the mid-1980s, pigs were selected for growth rate, feed efficiency, and carcass leanness using a simple individual selection index [START_REF] Ollivier | La loi sur l'élevage de 1966: Un bilan de 20 années de sélection rationnelle du porc en France[END_REF]. In 1985, a meat quality index, built as a predictor of the technological yield of cooked ham processing and computed as a linear combination of pH of the adductor femoris muscle, water-holding capacity, and reflectance of the biceps femoris muscle [START_REF] Jacquet | Prédiction du rendement technologique de la fabrication du «jambon de Paris» à l'aide de mesures prises à l'abattoir[END_REF], was introduced in the breeding goal. Candidates for selection were selected on a combined selection index based on their own performance and those of slaughtered siblings, with a restriction on meat quality index. At the end of the 1980s, a strong emphasis was placed on improving the litter size through the generalization of so-called "hyperprolific" breeding schemes. Finally, in the mid-1990s, standard selection indexes were replaced by more accurate predictors of breeding values based on multiple-trait BLUP animal model methodology [START_REF] Tribout | Continuous genetic evaluation of on farm and station tested pigs for production and reproduction traits in France[END_REF]. Management and other environmental conditions have improved progressively over the 21-yr period considered, with, for instance, an increasing knowledge of nutritional requirements of the growing pig and the generalization of AI.

Animals and Data Recording

First-Generation Animals and Phenotypes.

The design of the experiment is shown schematically in Figure 1. Two groups of LW pigs were produced in one of the INRA GEPA experimental units (Le Magneraud, Surgères, referred to as Le Magneraud hereafter) by inseminating 104 contemporary LW sows with frozen semen from LW boars born in 1977 (S77 sires) or with fresh semen of boars born in 1998 (S98 sires). Both S77 and S98 sires were chosen so as to be minimally related and as representative as possible of the population of AI boars in 1977 and 1998, respectively. Moreover, S77 and S98 boars were chosen to have a similar average superiority (i.e., a similar difference in average breeding value) for production traits over their contemporaries. Sows were randomly inseminated with either S77 or S98 semen, but particular attention was paid to having the same average parity number for each group of sows and to avoid inbreeding. Boars were all genotyped for the ryanodine receptor locus responsible for halothane sensitivity (HAL locus). All S98 boar and female founders were found to be free from the halothane sensitivity allele (Hal S ), but 3 of the 17 S77 sires were heterozygous at this same locus. Because studies on the frequency of halothane-sensitive pigs carried out in 1975 and 1976 had shown that the LW population could be regarded as free from the Hal S allele [START_REF] Ollivier | Fréquence du syndrome d'hyperthermie maligne dans des populations porcines françaises: Relation avec le développement musculaire[END_REF], the heterozygous progeny of these 3 boars was not considered representative of the 1977 LW population. As a consequence, all progeny of these 3 boars were also genotyped for the HAL locus, and heterozygous pigs were removed from the study (see discussion section). All experimental pigs considered in this study were hence negative for the HAL locus. A total of 30 litters from 17 S77 sires and 33 litters from 23 S98 sires (L77 and L98 litters, respectively) were produced. The sow herd was managed under a batch farrowing system, with a 3-wk interval between consecutive batches. These batches then became postweaning and performance-test batches of their progeny. Male L77 and L98 piglets were not castrated. At weaning (4 wk of age), one-half of the male and female piglets from each litter were randomly sampled and transferred to the INRA experimental unit at Bourges (Avord, referred to as Bourges hereafter), with the other one-half remaining in Le Magneraud unit.

In both herds, L77 and L98 male and female piglets were raised in pens of 12 animals and fed ad libitum from 10 until 22 wk of age. Animals were individually weighed at 10, 14, 18, and 22 wk of age and measured for backfat thickness at 14, 18, and 22 wk of age at 6 locations (on each side of the spine, 4 cm from the middorsal line at the shoulder, the last rib, and the hip joint, respectively) using a real-time ultrasound Aloka SSD-500 device (Ecotro Aloka, Tokyo, Japan).

Second-Generation Animals and Phenotypes. After puberty, 90 L77 females, 90 L98 females, 15 L77 males, and 15 L98 males were randomly chosen from a maximum number of litters and kept to produce a second generation of animals. Matings were performed within each experimental group (L77 and L98), and each female was kept to produce 5 successive litters to estimate realized genetic trends for reproduction and maternal ability traits (results not presented here). The piglets born in these litters are referred to hereafter as G77 and G98 animals. To disentangle direct and maternal effects on early piglet growth, cross-fostering was practiced within the first hours after birth. The objective was to have an equivalent number of G77 and G98 piglets raised by either L77 or L98 nursing sows and to standardize litters to either 7 or 13 piglets. This objective was difficult to achieve in practice, so the number of piglets ranged from 5 to 19, with 2 peaks around 7 and 13. Similarly, the proportions of G77 and G98 piglets in each litter were, on average, close to 50%, but with extreme values of 0 and 100%.

All male piglets were castrated within the first week after birth. At weaning (4 wk of age), a random sample of 298 piglets born in Le Magneraud was transferred to the other GEPA experimental herd (Rouillé, referred to as Rouillé hereafter). Moreover, 63 randomly sampled piglets born in the Bourges unit and 119 randomly sampled piglets born in Le Magneraud unit were sent to a fourth INRA experimental unit (Le Rheu, referred to as Le Rheu hereafter). All remaining pigs were fattened in their herd of origin (Le Magneraud or Bourges). In the 4 herds, animals were raised in discontinuous batches (each batch including G77 and G98 individuals of both sexes) and grouped in pens of 12 animals (except in Le Rheu, where each pen was made up of 2 animals) of the same sex and experimental group. They were given ad libitum access to water and to a standard pelleted diet formulated to contain 3,200 kcal of DE/kg and 17% CP from 10 wk of age until the day before slaughter, considered as the end of the test period. The pens in the Rouillé herd were equipped with Acema 64 electronic feeders (Acemo, Pontivy, France), allowing the recording of individual food consumption [START_REF] Labroue | Alimentation électronique dans les stations publique de contrôle des performances: paramètres descriptifs du comportement alimentaire[END_REF]. Slaughters occurred once per week on a fixed day, when pig BW reached approximately 105 kg [the average BW at the end of the test period was 106 kg (SD = 6.1 kg)].

Pigs were individually weighed at 10 and 20 wk of age and also the day before slaughter (23rd wk of age, on average, varying from 135 to 199 d of age in both the G77 and G98 groups). Backfat thickness was ultrasonically measured at 20 wk of age at the same 6 locations as for L77 and L98 animals. At the end of the test period, 2 samples of 120 animals fattened in the Bourges and Rouillé herds, balanced for experimental group and sex, were slaughtered in the INRA experimental slaughterhouses at Jouy-en-Josas and Saint-Gilles, respectively. The remaining pigs were slaughtered in different commercial slaughterhouses [i.e., Montfort-sur-Meu (Cooperl-Hunaudaye, Montfort-sur-Meu, France), Celles-sur-Belle (Socopa, Celles sur Belle, France), and Orléans Viande (Fleury les Aubrais, France) for pigs from the Le Rheu, GEPA, and Bourges units, respectively]. The same measurement protocol was applied in the different slaughterhouses. Carcass weight was recorded after evisceration on the day of slaughter. The day after slaughter, the length of the carcass from the pubis to the atlas, as well as the backfat thickness at the shoulder, last rib, and hip joint at the sectioned edge of the carcass were recorded. After chilling, the medial and dorsal faces of the right side of the hung carcass were photographed, following the protocol described by [START_REF] Laville | Prediction of composition traits of young Charolais bull carcasses using a morphometric method[END_REF] for carcass shape comparison purposes (pigs slaughtered at Montfort-sur-Meu, Jouy-en-Josas, and Saint-Gilles only). Additional measurements used for carcass grading [i.e., backfat thickness between the third and fourth lumbar vertebrae (G1) and between the third and fourth last ribs (G2), as well as loin eye depth between the third and fourth last ribs (M2)] were taken using a CGM probe (Sydel, Lorient, France; [START_REF] Daumas | Les méthodes de classement des carcasses de porc autorisées en France en 1997[END_REF]. Finally, a standardized cutting of the right half carcass was performed (Ministère de l'Agriculture et de la Forêt, 1990). Ham, loin, backfat, shoulder, belly, head, foot, leaf fat, and diaphragm were weighed. An additional measurement (i.e., loin eye area) was determined from the outline of the loin eye muscle at the seventh rib (traced on tracing paper). The number of pigs measured per herd and experimental group, as well as the number of fattening batches and slaughter groups per herd are shown in Table 1.

Power of the Design

The power of the actual design (i.e., the probability of detecting a trend if it exists) can be computed as the power of a 2-sided test (i.e., hypothesis 1, μ 98 ≠ μ 77 vs. null hypothesis, μ 98 = μ 77 ). If the test statistic T is nor-mally distributed with variance σ 2 and mean 0 under the null hypothesis and (μ 98 -μ 77 ) under hypothesis 1, the power can be computed as

Pr ( ) Pr Pr ( / ) T T U z U c > é ë ê ù û ú = < - - é ë ê ê ê ù û ú ú ú + > - - a m m s m m a 77 98 1 2 77 98 8 1 2 s a + é ë ê ê ê ù û ú ú ú - z ( / ) ,
where α is the significance level of the test,

T c (α) is the α-level critical value, U = (T -μ 98 -μ 77 )/σ ~N(0, 1) and z (1-α/2) is the (1 -α/2)-level critical value of U.
Ignoring covariances between experimental groups attributable to female founders, the variance σ 2 is the sum of the sampling variance of the mean of each experimental group, calculated as [START_REF] Smith | A note on the efficiency of different forms of comparison among foreign breeds[END_REF], where s is the number of sires, d is the number of dams per sire, n is the number of offspring per dam, and t 1 and t 2 are, respectively, the phenotypic correlations between full-sibs (t 1 = h 2 /2 + c 2 , where h 2 is the heritability of the trait and c 2 is the proportion of phenotypic variance from birth litter environmental origin), and halfsibs (t 2 = h 2 /4) [START_REF] Falconer | Introduction to Quantitative Genetics[END_REF]. Four cases were considered, according to the size of the G77 and G98 samples and their familial structure in the present design: A) nds = 1,000 pigs, ds = 75 dams, and s = 18 boars; B) nds = 500 animals, ds = 75 dams, and s = 18 boars; C) nds = 180 animals, ds = 56 dams, and s = 18 boars; D) nds = 120 animals, ds = 48 dams, and s = 17 boars. Three heritabilities (h 2 = 0.2, 0.5, and 0.7), corresponding to the range of heritability values for the traits investigated, were considered for each of the 4 cases. The power of the corresponding designs is shown in Figure 2, assuming a value of 5% for α.

V n t n d t nds = + - + - é ë ê ù û ú 1 1 1 1 2 ( ) ( ) (

Traits Analyzed

Growth traits analyzed included ADG from the beginning to the end of the test period (i.e., from 10 to 22 wk of age in first-generation pigs, and from 10 wk of age to slaughter in second-generation pigs), as well as between successive BW measurements (i.e., from 10 to 14 wk, 14 to 18 wk, and 18 to 22 wk of age in firstgeneration pigs, and from 10 to 20 wk of age in secondgeneration pigs). Body weight at 20 wk of age was not measured on first-generation pigs, but was derived by interpolating BW at 18 and 22 wk of age to compute ADG from 10 to 20 wk of age for L77 and L98 pigs.

Individual cumulative feed consumption was computed as the sum of the amounts of food ingested by an animal at each of its feed intakes during the whole test period (10 to 20 wk of age) and divided by either the BW increase or the length of the test period to compute feed conversion ratio (FCR) and ADFI, respectively.

Carcass composition traits included average backfat thickness (computed as the mean of the 6 ultrasonic measurements) at 14, 18, and 22 wk of age in generation 1 and at 20 wk of age in generation 2; dressing percentage; pubis-atlas length; backfat thickness measured at the sectioned edge of the carcass at the levels of the shoulder, the last rib, and the hip joint; G1 and G2 fat depth; M2 loin eye depth; and loin eye area as well as ham, loin, backfat, shoulder, belly, head, feet, leaf fat, and diaphragm weights. An estimated carcass lean content was computed from primal cut weights, expressed as a percentage of half-carcass weight [START_REF] Métayer | Estimation par découpe de la teneur en viande maigre des carcasses de porc[END_REF]:

Estimated carcass lean content = 5.684 + 1.197% ham + 1.076% loin -1.059% backfat.

To investigate changes in carcass shape, 10 measurements were taken on carcass numeric images (Figure 3) using Optimas image analysis software (Media Cibernetics, Silver Spring, MD). On the dorsal face, measurements included back dorsal width at the pelvis and 2 angles reflecting the thickness of ham muscles (i.e., the angle between the vertical and a line set down the medial hind leg plumb and the angle between the vertical and a line set down the lateral hind leg plumb). On The number of records varies for the different traits.

2

First generation = offspring of boars born in 1977 (L77 animals) and in 1998 (L98 animals).

3

Second generation = offspring of L77 boars and L77 females (G77 animals) and of L98 boars and L98 females (G98 animals).

the medial face, measurements included a third angle related to the ham muscle thickness (i.e., the angle between the vertical and a line set down the posterior hind leg plumb), 2 indicators of hind leg length (i.e., the distances between the calcaneal tip and the cranial edge of the pubic symphysis and between insertion of the tendon on the toes and the cranial edge of the pubic symphysis). The length of the back was assessed by 3 measurements (i.e., the distances between the cranial edge of the pubic symphysis and the last lumbar vertebra, between the last lumbar vertebra and the last thoracic vertebra, and between the last thoracic vertebra and the first thoracic vertebra). Finally, the depth of the thoracic cavity was estimated by the dorso-ventral height at the level of the thorax. Descriptive statistics for the 44 traits analyzed are presented in Supplemental Table 1 (http://jas.fass.org/content/vol88/issue9/).

Statistical Analyses

The data were analyzed using a mixed linear model, taking into account relationships within each of the 2 groups of pigs. With the exception of ADG from 10 to 20 wk of age, traits differed between generations and were consequently analyzed on a within-generation basis. The model used for each trait was determined using a 2-step procedure. First, the fixed effects and covariates to be included in the final analyses were determined using a fixed linear model with the GLM procedure (SAS Inst. Inc., Cary, NC). The fixed effects investigated were the experimental group (L77 vs. L98, or G77 vs. G98 pigs), the sex (males or females in the first generation; castrates or females in the second generation), the fattening batch (defined as the pigs born during the same week and fattened in the same building), and the abattoir (for traits measured after slaughter). The covariates tested were BW at 10 wk of age (for ADG from 10 to 14 wk of age in first-generation pigs, ADG from 10 to 20 wk of age in both generations of pigs, ADG from 10 wk of age to slaughter in secondgeneration pigs, FCR, and ADFI), at 14 wk of age (for ADG from 14 to 18 wk of age and ultrasonic backfat thickness at 14 wk of age in first-generation pigs), at 18 wk of age (for ADG from 18 to 22 wk of age and ultrasonic backfat thickness at 18 wk of age in firstgeneration pigs), at 20 wk of age (for ultrasonic backfat thickness at 20 wk of age in second-generation pigs), at 22 wk of age (for ultrasonic backfat thickness at 22 wk of age in first-generation pigs), and at the end of the test period (for FCR, ADFI, and all the traits mea- sured after slaughter). The effects of cross-fostering, of the number of piglets nursed by the nursing sow, and interactions between fixed effects as well as between fixed effects and covariates were tested in preliminary analyses. With very few exceptions, they were found to be nonsignificant (P > 0.15) and had no effect on estimates of contrasts between genetic types. They were consequently removed from the final analyses. A joint analysis of the 2 generations was also performed for ADG from 10 to 20 wk of age by adding a generation effect and its interaction with the experimental group in the model. Once the fixed part of the model was established, it was included in a mixed linear model including the additive genetic value of each animal and the common effect of birth litter as random effects. The fraction of the phenotypic variance attributable to random effects was then estimated using REML methodology applied to the single-trait individual animal model described above with VCE software [START_REF] Neumaier | Restricted maximum likelihood estimation of covariances in sparse linear models[END_REF]. The effects considered and the estimates of heritability and common litter effect (when significant) for each trait analyzed are given in Supplemental Table 2 (http://jas.fass.org/content/vol88/ issue9/). These REML estimates were then introduced as priors in the same mixed models to compute best linear unbiased estimates of the contrasts between genetic groups and to test their significance. The analyses were performed with PEST software [START_REF] Groeneveld | PEST, a general purpose BLUP package for multivariate prediction and estimation[END_REF]. Finally, contrasts between genetic groups (D 98-77 ) and their SE (seD 98-77 ) were used to estimate genetic trends from 1977 to 1998 (ΔG) and their SE [START_REF] Smith | A note on the efficiency of different forms of comparison among foreign breeds[END_REF] as ΔG = (2 × D 98-77 ) and SE = (2 × seD 98-77 ).

RESULTS

Performance Traits

Results for growth traits are given in Table 2. Positive estimates of genetic trends for ADG over the whole test period were obtained in both generations, but with a smaller and nonsignificant (P ≥ 0.15) estimate in the first generation (+45 ± 31 g/d for ADG from 10 to 22 wk of age) and a larger, highly significant (P = 0.0043) estimate in second-generation pigs (+77 ± 27 g/d for ADG from 10 wk of age to slaughter). The improvement mainly occurred during late growth: no significant trend was obtained in first-generation pigs from 10 to 14 wk or from 14 to 18 wk of age, whereas ADG significantly increased over the period from 18 to 22 wk of age in first-generation pigs (+116 ± 56 g/d for ADG from 18 to 22 wk of age). Similarly, the contrast between G77 and G98 animals did not reach significance (P ≥ 0.14) for ADG from 10 to 20 wk of age but became significant (P = 0.0043) when considering the whole test period (ADG from 10 wk of age to slaughter in second-generation pigs). Finally, the joint analysis of ADG from 10 to 20 wk of age in second-generation pigs did not reveal any difference between estimated genetic trends in the 2 generations, with a significance level for the generation × experimental group interaction test of P = 0.75 (results not presented).

Although only measured on a limited subsample of second-generation pigs, estimated genetic trends for ADFI and FCR showed a tendency toward an increase (P = 0.09) in ADFI (0.16 ± 0.10 kg) and a significant (P = 0.006) improvement in FCR (-0.30 ± 0.11 kg/ kg). The estimated genetic trends for ultrasonic backfat thickness are given in Table 3. They showed a strong and consistent decrease in carcass adiposity (all P-values <0.0001) in both generations. The relative reduction in backfat thickness tended to increase with age [-2.7 phenotypic SD (σ ph ) and -2.8 σ ph at 18 and 22 wk of age, respectively, vs. -1.9 σ ph at 14 wk of age]. It also varied according to the measurement site (Figure 4), with a stronger decrease at the level of the hip joint (-5.9 mm, or -2.7 σ ph ) and of the last rib (-4.9 mm, or -2.6 σ ph ) than at the shoulder level (-4.7, or -1.9 σ ph ).

Carcass Traits

Estimated genetic trends for carcass traits are shown in Tables 4 and5. Selection did not significantly affect dressing percentage or carcass length (P > 0.2).

Similarly, none of the 10 measurements of carcass shape showed any significant change between 1977 and 1998 (P > 0.3; Table 5). Conversely, results showed a highly significant increase in loin thickness (loin eye depth and loin eye area; P < 0.006). Estimated genetic trends for backfat thickness measured on the sectioned edge of the carcass showed a highly significant decrease (P < 0.001). They were consistent with the estimated trends for ultrasonic backfat measurements, with stronger reductions at the hip joint (-7.9 mm, or -2.0 σ ph ) and last rib (-7.3 mm, or -2.0 σ ph ) than at the shoulder (-7.0 mm, or -1.4 σ ph ). However, it should be emphasized that the relative increases in loin eye depth and Probability associated with the null hypothesis (H0): ΔG = 0 (P-value) for each trait.

7

Results obtained by analyzing the data of the first and second generations independently.

8

Second generation = offpring of L77 boars and L77 sows (G77 animals) and offspring of L98 boars and L98 sows (G98 animals). FCR = feed conversion ratio. Probability associated with the null hypothesis (H0): ΔG = 0 (P-value) for each trait.

loin eye area were much less (1.1 and 1.2 σ ph , respectively) than the decrease in backfat thickness.

The estimated genetic trend pattern was similar when considering primal cut weights. An important decrease in backfat thickness and a strong increase in loin and ham weights were obtained, resulting in a large improvement in estimated carcass lean content (+8.6 percentage points, or 2.5 σ ph ). When expressed in SD units, the improvement in backfat weight (-2.4 σ ph ) was twice as large as that of loin and ham weights (1.2 and 1.1 σ ph , respectively). Selection also affected internal fat with a significant reduction in leaf fat weight 1 BTSEC = backfat thickness measured at the sectioned edge of the carcass; G1 = backfat thickness at the third and fourth lumbar vertebrae; G2 = backfat thickness at the third and fourth last ribs; M2 = loin eye depth at the third and fourth last ribs; ECLC = estimated carcass lean content.

2 Number of observations per population.

3 Average performance of phenotyped animals. 4 REML estimate of phenotypic SD. 5 Estimated realized genetic trend from 1977 to 1998 (ΔG) and its SE. 6 Probability associated with the null hypothesis (H0): ΔG = 0 (P-value) for each trait. 7 Second-generation intercross of boars born in 1977 and base females.

8 Second-generation intercross of boars born in 1998 and base females. 9 CGM (Sydel, Lorient, France).

(-0.33 kg, or -1.2 σ ph ; P = 0.02), tended to reduce belly weight (P = 0.07) and to increase feet weight (P = 0.08). Conversely, no significant trend was observed for head and shoulder weights.

DISCUSSION

The use of frozen semen is a simple and efficient way to estimate realized genetic trends in a population [START_REF] Smith | A note on the efficiency of different forms of comparison among foreign breeds[END_REF]. Indeed, it allows genetic trends to be estimated on a large number of traits that are not routinely recorded. This is particularly attractive for phenotypes that are difficult or expensive to measure on a large number of animals, such as quality, behavioral, or physiological components of traits of interest. Genetic trends can also be estimated in various environments (e.g., feeding regimens) to check for potential genotype × environment interactions. Finally, the estimated genetic trends are basically contrasts between the 2 levels of a fixed effect in a linear model, and are therefore much more robust than annual averages of BLUP EBV, which are known to be sensitive to the data modeling and to the genetic parameters used as priors. Because some of the animals tested in the present design were related (siblings, offspring of related parents), it was decided to model the data with an animal model to take these relationships into account and not to overestimate the significance of the estimated differences between the offspring of S77 and S98 boars. This does not imply that the results are prior dependent because best linear unbiased estimates are more robust than BLUP to bad priors. Indeed, a sensitivity analysis of our results performed by increasing and decreasing the heritabilities used in PEST by 30% gave results that remained unchanged.

In spite of its interest, frozen semen has seldom been used to estimate genetic trends in livestock. In pigs, besides the above-mentioned French studies (Molénat et al., 1986;[START_REF] Ollivier | Progrès génétique des porcs Large White et Landrace français de 1977 à 1987[END_REF], the only study was that of [START_REF] Schwab | Deposition rates and accretion patterns of intramuscular fat, loin muscle area, and backfat of Duroc pigs sired by boars from two time periods[END_REF], who estimated genetic trends from 1985 to 2002 for growth, backfat, loin muscle area, and intramuscular fat in American Durocs. This situation is presumably because such an experiment is rather complex to set up. Indeed, semen has to be collected on a random sample of males many years before the implementation of the design, which requires long-term planning of the experiment. Substantial financial support is also required for semen collection and long-term storage, and to compensate for the reduced conception rates attributable to the use of frozen semen, the reduced commercial value of unselected animals, and the measurement costs of numerous traits of interest.

Another limitation of frozen semen designs is that, by construction, they allow comparison of the population genetic levels only at the beginning and at the end of the period considered, without providing the actual shape of the genetic trends. It is rather likely that trends have not been linear in French LW because the relatively simple breeding goal from the late 1970s based on growth, feed efficiency, and carcass merit [START_REF] Ollivier | La loi sur l'élevage de 1966: Un bilan de 20 années de sélection rationnelle du porc en France[END_REF] has progressively become more complex with the inclusion of meat quality since the mid-1980s and, above all, the emergence of litter size at birth as the major trait of interest in dam populations since the early 1990s. Selection on these new traits has reduced selection pressure on the formerly selected traits, so 1 LONJAM1 = hind leg length between insertion of tendon on toes and the cranial edge of pubic symphysis; LONJAM2 = hind leg length between the calcaneal tip and the cranial edge of pubic symphysis; SYMP = distance between cranial edge of pubic symphysis and the last lumbar vertebra; LOMB = distance between the last lumbar vertebra and the last thoracic vertebra; THOR = distance between the last thoracic vertebra and the first thoracic vertebra; PROFPOIT = dorso-ventral height at thorax; ANG1 = angle between the vertical and a line set down the posterior hind leg plumb; ANGINT = angle between the vertical and a line set down the medial hind leg plumb; ANGEXT = angle between the vertical and a line set down the lateral hind leg plumb; LARGBA = back dorsal width at pelvis. trends computed over the 21 yr are inevitably less than those computed over a shorter period of time corresponding to a single breeding goal. This at least partly explains the smaller yearly trends obtained in the current study as compared with those previously obtained in the same population by [START_REF] Molénat | Utilisation de la semence congelée pour l'estimation du progrès génétique sur les caractères de production chez le porc[END_REF] and [START_REF] Ollivier | Progrès génétique des porcs Large White et Landrace français de 1977 à 1987[END_REF]Ollivier et al. ( ) over the 1977Ollivier et al. ( to 1982Ollivier et al. ( and 1977Ollivier et al. ( to 1987 time periods, respectively (the most striking difference being for ADG, with yearly trends of 24.5 ± 5.9 g/d and 12.8 ± 3.2 g/d, respectively, vs. 3.7 ± 1.3 g/d in the current study). Moreover, the studies by [START_REF] Molénat | Utilisation de la semence congelée pour l'estimation du progrès génétique sur les caractères de production chez le porc[END_REF] and [START_REF] Ollivier | Progrès génétique des porcs Large White et Landrace français de 1977 à 1987[END_REF] were conducted on smaller numbers of animals (203 and 269 animals, respectively), so the limited accuracy of their estimates of genetic trends can also explain the differences from current results.

The fact that animals are generally compared under current management conditions that would favor modern genotypes and lead to biased results is often put forward as another limitation of frozen semen designs. The latter can address genotype × environment interactions by comparing experimental groups under different management conditions, but at the expense of an increasing size of the design. Investigation of the genotype × feeding regimen interaction was one of the initial goals of the project, but this could not be addressed because of both financial and experimental limitations.

Contrary to the above-mentioned experiments, the current study also aimed at estimating genetic trends for reproductive traits, which offered the opportunity to set up a 2-generation design. This extension of the design had several advantages. First, it allowed the size and, consequently, the power of the experiment to be increased. Second, the potential bias attributable to the smaller size of the litters resulting from frozen semen no longer exists in second-generation pigs. Finally, the S77 boars and the "modern" sows can be considered as belonging to 2 different populations, given the large number of generations separating them; consequently, the potential heterosis effects that could have appeared when mating S77 boars to "modern" sows would be divided by 2 in the second generation of the design [START_REF] Falconer | Introduction to Quantitative Genetics[END_REF]. This latter hypothesis could be addressed for only 1 trait (i.e., ADG from 10 to 20 wk of age), which is one of the most likely to exhibit heterosis effects. The lack of a generation × genotype interaction suggests that heterosis effects are absent, or at least very limited, in first-generation pigs.

As mentioned above, 3 of the 17 S77 sires were found heterozygous at the ryanodine receptor locus. This was unexpected because tests for malignant hyperthermia syndrome susceptibility performed on samples of the LW population in 1975 and 1976 showed that the French LW population could be considered free from the Hal S allele [START_REF] Ollivier | Fréquence du syndrome d'hyperthermie maligne dans des populations porcines françaises: Relation avec le développement musculaire[END_REF]. The presence of 3 heterozygous boars in the S77 sample showed that the Hal S allele was still present in the population in the late 1970s, but at a very low frequency, and was due to a greater probability of heterozygotes being selected, given the strong favorable effects of HAL on carcass lean content. It has to be added that the use of AI was minimal at the end of the 1970s, so the diffusion of the Hal S allele on a large scale was limited, as shown by the results of [START_REF] Ollivier | Fréquence du syndrome d'hyperthermie maligne dans des populations porcines françaises: Relation avec le développement musculaire[END_REF]. As a consequence, it was decided that all progeny of the 3 heterozygous S77 boars be removed from the study, to have a sample of pigs that was as representative as possible of the 1977 French LW population. The consequences of these eliminations on the estimated genetic trends were nevertheless very small because fewer than 7% of the piglets produced were likely to be heterozygous for the ryanodine receptor locus.

The significant, favorable trends obtained in the present study for growth rate, feed efficiency, and carcass composition tended to be larger than those previously obtained in the same population by [START_REF] Tixier | Estimated genetic trends for growth and carcass traits in two French pig breeds[END_REF]) between 1970[START_REF] Falconer | Introduction to Quantitative Genetics[END_REF][START_REF] Ducos | Utilisation d'un modèle animal multi-caractères pour estimer l'évolution génétique de six caractères de croissance, carcasse et qualité de la viande entre 1977 et 1990, dans les races porcines Large White et Landrace français[END_REF]) between 1977and 1990. Indeed, yearly trends for ADG, backfat thickness, and FCR amounted to, respectively, 4.3, -9.0 to -13.3 (depending on the measurement period), and -6.5% of the σ ph in the current study vs. 3.5, -13.7, and -4.6% of the σ ph by [START_REF] Tixier | Estimated genetic trends for growth and carcass traits in two French pig breeds[END_REF] and 0.8, -6.2, and -3.6% of the σ ph by [START_REF] Ducos | Utilisation d'un modèle animal multi-caractères pour estimer l'évolution génétique de six caractères de croissance, carcasse et qualité de la viande entre 1977 et 1990, dans les races porcines Large White et Landrace français[END_REF]. Both studies estimated genetic trends using mixed model methodology, but used only data from test stations without considering on-farm data, which may have resulted in some underestimation of actual genetic trends.

Yearly trends are also rather comparable with those obtained in other populations. Indeed, similar moderate trends were obtained for growth rate by [START_REF] Kennedy | Genetic changes in Canadian performance-tested pigs for fat depth and growth rate[END_REF] on 4 Canadian breeds (2.9 to 4.1% of the σ ph for days to 100 kg) and by [START_REF] Chen | Genetic parameters and trends for lean growth rate and its components in U.S. Yorkshire, Duroc, Hampshire, and Landrace pigs[END_REF] on American Yorkshires, Landraces, and Durocs (2.3, 3.1, and 3.2% of the σ ph for age at 113 kg, respectively). Trends for average backfat thickness were much larger and somewhat more variable between populations. Results from the current study were slightly larger than the estimates reported by [START_REF] Chen | Genetic parameters and trends for lean growth rate and its components in U.S. Yorkshire, Duroc, Hampshire, and Landrace pigs[END_REF] in 4 American breeds (-6.4 to -9.2% of the σ ph ) and by [START_REF] Kennedy | Genetic changes in Canadian performance-tested pigs for fat depth and growth rate[END_REF] in 4 Canadian breeds (-4.4 to -9.4% of the σ ph ). Differences in the amount of response to selection mean that more emphasis was placed on the reduction of backfat thickness and on the improvement of carcass lean content than on the improvement of growth rate. It should also be emphasized that estimates of genetic trends are generally less than the maximum improvement that is theoretically possible in the pig population (e.g., [START_REF] Smith | Rates of genetic change in farm livestock[END_REF]. Reasons for these discrepancies are numerous, but are at least partly related to the fact that breeding goals in pig dam lines include additional major objective traits that are independent of (litter size) or unfavorably related to (meat quality) performance traits and reduce the selection pressure on those traits.

This study also provides additional information on the way genetic improvement has acted in the French LW population. In particular, it should be emphasized that the effects of selection on growth and backfat deposition were much larger at the end of the test period. [START_REF] Schwab | Deposition rates and accretion patterns of intramuscular fat, loin muscle area, and backfat of Duroc pigs sired by boars from two time periods[END_REF] also reported differences in daily accretion of backfat between pigs sired by current vs. old-time period Duroc boars, but without any difference in growth rate. Numerous hypotheses can be proposed to explain these differences; the 2 populations may differ in terms of food intake, protein, and fat accretion curves, which would result in differences in the amount and the partitioning of energy to maintenance or to protein and fat deposition during the growth period. It may, for instance, be hypothesized that food intake is a limiting factor in the French LW breed at the beginning of the growth period, which might restrict genetic improvement during early growth. Such questions could be addressed using suitable growth models (e.g., [START_REF] Knap | Stochastic simulation of growth in pigs: Protein turn-over-dependent relations between body composition and maintenance requirements[END_REF][START_REF] Schinckel | Describing the pig[END_REF] but require specific experiments to estimate the parameters associated with these models.

Furthermore, selection has altered the distribution of lean and fat tissues, as well as loin muscle thickness, without any change in carcass dimensions (i.e., without affecting the size of the skeletal frame). The amounts of both backfat and internal fat tissues have been strongly reduced in favor of lean tissues in both the loin and the ham. The reduction has not been homogeneous all along the carcass because the reduction has been larger at the back of the carcass in spite of a large fat depth at the shoulder. This result is consistent with the result of [START_REF] Legault | Expérimentation sur le porc chinois en France. II-Performances de production en croisement avec les races européennes[END_REF], who showed in a breed comparison that the profile of the backfat layer differs between lean and fat genotypes. The gradient between fat depths at the levels of the rump and of the back is clearly positive in fat genotypes, such as the Meishan breed, but reduces in leaner genotypes and becomes negative in very lean breeds, such as the Pietrain. A reduction of intramuscular fat content can also be expected based on genetic correlation estimates with carcass composition traits (e.g., [START_REF] Sellier | Genetics of meat and carcass traits[END_REF] and on the results of [START_REF] Schwab | Deposition rates and accretion patterns of intramuscular fat, loin muscle area, and backfat of Duroc pigs sired by boars from two time periods[END_REF], who reported a decrease in intramuscular fat content when comparing current vs. old-time Duroc boars.

In conclusion, this study has demonstrated that the use of frozen semen is a useful practice for investigating the consequences of selection in animal populations. It has been shown that large improvements have been achieved in the French LW population for growth, feed efficiency, and carcass lean content. Additional studies are necessary to investigate the effects of selection on other economically important traits, such as reproduction or meat quality. The use of growth models would be useful to better understand the impact of selection on the different components of pig growth to optimize food utilization, lean tissue growth, and quality while minimizing the environmental impact of pig production.

Figure 1 .

 1 Figure 1. Experimental design. S77 = boars born in 1977; S98 = boars born in 1998; L77 and L98 = first generation of the design; G77 and G98 = second generation of the design; LW = French Large White.

Figure 2 .

 2 Figure 2. Power of the design for different sample sizes and heritability (h 2 ) values, as a function of the realized genetic trend from 1977 to 1998 (in phenotypic SD units), assuming a 5% type I error. A) Offspring (n = 1,000) from 18 boars and 75 dams measured in each experimental group. B) Offspring (n = 500) from 18 boars and 75 dams measured in each experimental group. C) Offspring (n = 180) from 18 boars and 56 dams measured in each experimental group. D) Offspring (n = 120) from 17 boars and 48 dams measured in each experimental group

Figure 3 .

 3 Figure3. Carcass shape measurements: LONJAM1 = hind leg length between insertion of tendon on toes and the cranial edge of pubic symphysis; LONJAM2 = hind leg length between the calcaneal tip and the cranial edge of pubic symphysis; SYMP = distance between cranial edge of pubic symphysis and the last lumbar vertebra; LOMB = distance between the last lumbar vertebra and the last thoracic vertebra; THOR = distance between the last thoracic vertebra and the first thoracic vertebra; PROFPOIT = dorso-ventral height at thorax; ANG1 = angle between the vertical and a line set down the posterior hind leg plumb; ANGINT = angle between the vertical and a line set down the medial hind leg plumb; ANGEXT = angle between the vertical and a line set down the lateral hind leg plumb; LARGBA = back dorsal width at pelvic.

Figure 4 .

 4 Figure 4. Estimated genetic trends (±SE) for ultrasonic backfat thickness at the hip joint, last rib, and shoulder recorded at 14, 18, and 22 wk of age on first-generation pigs. A) Estimated genetic trends, expressed in millimeters. B) Estimated genetic trends, expressed in phenotypic SD (σ) units.

Table 1 .

 1 Numbers of pigs measured per herd and experimental group, and number of fattening batches and slaughter groups per herd in the first and second generations of the design 1

			First generation, 2 No.		Second generation, 3 No.
		Animals		Animals
				Fattening			Fattening	Slaughter
	Herd	L77	L98	batches	G77	G98	batches	groups
	Le Magneraud	88	95	7	228	450	13	12
	Bourges	100	119	7	886	1,027	24	54
	Rouillé				122	124	7	17
	Le Rheu				65	109	1	5
	Total	188	214	14	1,301	1,710	45	88
	1

Table 2 .

 2 Estimated genetic trends from 1977 to 1998 for ADG and feed efficiency

		Observations 2
				Mean			Pr > |f| for
	Trait 1	L77	L98	performance 3	σ 4	ΔG ± SE 5	H0: ΔG = 0 6
	ADG from 10 to 22 wk of age, g/d	187	214	872	94	45 ± 31	0.15
	ADG from 10 to 20 wk of age, 7 g/d	187	214	838	91	29 ± 30	0.34
	ADG from 10 to 14 wk of age, g/d	187	214	690	106	-15 ± 37	0.69
	ADG from 14 to 18 wk of age, g/d	188	214	870	129	17 ± 46	0.71
	ADG from 18 to 22 wk of age, g/d	188	214	1,032	170	116 ± 56	0.038
	Trait 8	G77	G98
	ADG from 10 wk of age to slaughter, g/d	1,287	1,610	883	85	77 ± 27	0.0043
	ADG from 10 to 20 wk of age, 7 g/d	988	1,213	858	103	36 ± 24	0.14
	FCR from 10 to 20 wk of age, kg/kg	104	117	2.76	0.22	-0.30 ± 0.11	0.006
	ADFI from 10 to 20 wk of age, kg	104	117	2.07	0.17	0.16 ± 0.10	0.09
	1 First generation = offpring of boars born in 1977 (L77 animals) and offspring of boars born in 1998 (L98 animals).
	2 Number of L77 and L98 (first-generation) or G77 and G98 (second-generation) measurements.
	3 Average performance of phenotyped animals.
	4 REML estimate of phenotypic SD.

5

Estimated realized genetic trend from 1977 to 1998 (ΔG) and its SE. 6

Table 3 .

 3 Estimated genetic trends from 1977 to 1998 for ultrasonic backfat thickness (UBT)

		Observations 2
				Mean			Pr > |f| for
	Trait 1	L77	L98	performance 3	σ 4	ΔG ± SE 5	H0: ΔG = 0 6
	Average UBT at 14 wk of age, mm	188	214	9.0	1.0	-1.9 ± 0.4	<0.0001
	Average UBT at 18 wk of age, mm	187	214	11.1	1.5	-4.1 ± 0.6	<0.0001
	Average UBT at 22 wk of age, mm	188	212	14.8	2.0	-5.6 ± 0.8	<0.0001
	Trait 7	G77	G98
	UBT at 20 wk of age at hip joint, mm	988	1,213	12.9	2.20	-5.9 ± 0.9	<0.0001
	UBT at 20 wk of age at last rib, mm	988	1,213	10.7	1.86	-4.9 ± 0.7	<0.0001
	UBT at 20 wk of age at shoulder, mm	988	1,213	17.0	2.42	-4.7 ± 0.8	<0.0001
	Average UBT at 20 wk of age, mm	988	1,213	13.5	1.86	-5.2 ± 0.7	<0.0001

1 First generation = offpring of boars born in 1977 (L77 animals) and offspring of boars born in 1998 (L98 animals). 2 Number of L77 and L98 (first-generation) or G77 and G98 (second-generation) measurements. 3 Average performance of phenotyped animals. 4 REML estimate of phenotypic SD. 5 Estimated realized genetic trend from 1977 to 1998 (ΔG) and its SE.

6

Table 4 .

 4 Estimated genetic trends from 1977 to 1998 for carcass traits

		Observations 2
				Mean			Pr > |f| for
	Trait 1	G77 7	G98 8	performance 3	σ 4	ΔG ± SE 5	H0: ΔG = 0 6
	Dressing percentage	852	996	77.6	1.80	-0.7 ± 0.6	0.26
	Pubis-atlas length, mm	579	673	991.9	27.1	-6.6 ± 10.8	0.54
	BTSEC, mm
	At hip joint	553	649	18.6	3.9	-7.9 ± 1.6	<0.0001
	At last rib	553	649	19.2	3.3	-7.3 ± 1.2	<0.0001
	At shoulder	553	649	30.6	4.6	-7.0 ± 1.5	<0.0001
	CGM 9 measurements, mm
	G1	857	1,032	19.7	3.8	-10.0 ± 1.5	<0.0001
	G2	857	1,032	17.0	3.5	-8.9 ± 1.3	<0.0001
	M2	857	1,032	52.1	5.4	6.0 ± 2.1	0.0054
	Loin eye area at 7th rib, cm 2	170	167	43.3	5.4	6.6 ± 2.1	0.002
	Primal cut wt, kg
	Ham	506	559	9.67	0.50	0.56 ± 0.20	0.005
	Loin	504	560	10.45	0.68	0.85 ± 0.30	0.004
	Shoulder	506	559	8.32	0.51	-0.17 ± 0.16	0.27
	Backfat	504	559	3.04	0.60	-1.41 ± 0.28	<0.0001
	Belly	506	560	5.30	0.45	-0.25 ± 0.14	0.07
	Feet	482	549	0.97	0.07	0.05 ± 0.03	0.08
	Head	320	403	4.72	0.34	0.18 ± 0.14	0.18
	Leaf fat	116	120	0.96	0.27	-0.33 ± 0.14	0.02
	Diaphragm	116	120	0.31	0.03	0.03 ± 0.02	0.06
	ECLC, %	481	550	55.7	3.5	8.6 ± 1.7	<0.0001

Table 5 .

 5 Estimated realized genetic trends from 1977 to 1998 for carcass shape measurements

		Observations 2
				Mean			Pr > |f| for
	Trait 1	G77 7	G98 8	performance 3	σ 4	ΔG ± SE 5	H0: ΔG = 0 6
	LONJAM1, cm	151	146	48.8	2.1	0.86 ± 0.98	0.38
	LONJAM2, cm	156	158	31.9	1.3	0.44 ± 0.62	0.48
	SYMP, cm	156	158	10.9	1.0	-0.10 ± 0.30	0.71
	LOMB, cm	156	158	28.0	1.6	-0.08 ± 0.68	0.90
	THOR, cm	156	158	47.9	2.5	1.32 ± 1.26	0.30
	PROFPOIT, cm	156	158	31.7	1.4	0.72 ± 0.70	0.31
	ANG1, degrees	156	158	50.4	4.6	-1.40 ± 2.02	0.49
	ANGINT, degrees	100	79	46.1	6.2	-2.10 ± 3.62	0.56
	ANGEXT, degrees	100	79	24.9	3.3	0.88 ± 1.42	0.53
	LARGBA, cm	100	79	20.0	0.8	0.12 ± 0.38	0.74

  2 Number of observations per population. 3 Average performance of phenotyped animals. 4 REML estimate of phenotypic SD. 5 Estimated realized genetic trend from 1977 to 1998 (ΔG) and its SE. 6 Probability associated to the null hypothesis (H0): ΔG = 0 (P-value) for each trait. 7 Second-generation intercross of boars born in 1977 and base females. 8 Second-generation intercross of boars born in 1998 and base females.

The financial support of the "Innovative Actions" from the French Ministry of Agriculture is gratefully acknowledged.

The authors thank the staff of INRA experimental herds and laboratories implied in this experiment, in particular Francis Bussières from the INRA Experimental Unit for Caprine and Porcine Insemination in Rouillé for collecting, conditioning, and taking care of the semen of 1977 boars for more than 20 yr.

3 Corresponding author: thierry.tribout@jouy.inra.fr ReceivedJuly 31, 2009. Accepted May 17, 2010.

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