Final report to Minnesota Pork Board Funded by the Pork Checkoff

Effects of phase feeding gestating and lactating sows on reproduction performance, piglet robustness at birth and post-weaning

Samuel Kofi Baidoo, Associate Professor Southern research and Outreach Center, University of Minnesota, Waseca, MN 56093

Abstract

Effect of phase feeding versus conventional feeding was investigated on sow reproductive performance for two successive parities. A total of 240 mixed parity (1-7) sows were assigned to two dietary treatments on the week of insemination. Sows were blocked by parity and had similar body weight (BW) and backfat (BF). The phase-fed sows (n = 121, BW = 227.0 ± 3.6 kg, BF = 16.5 ± 0.7 mm) received 0.4%, 0.57% and 0.7 % SID lysine diets in three different periods 1(d0 (breeding) - d35), 2 (d35- d70) and 3 (d70 – d109), respectively during gestation. The control sows (n = 120, BW = 227.7 ± 3.5 kg, BF =16.3 ± 0.7 mm) were fed a 0.57% SID lysine diet throughout gestation. Sows were moved to farrowing crates on d109 and assigned two different feeding regimens in a 2 x 2 factorial arrangement. Sows were fed a conventional lactation diet (1.0% standard ileal digestible (SID) lysine throughout lactation throughout lactation or test diets (SID lysine 0.8%, 1.0%, and 1.2%) for phases 1(d 0-6), 2(d 7-12) and 3 (d 13 - weaning) respectively. The sows on phase feeding regimens and on control regimen received control diets during successive gestation and lactation periods. Ratios of lysine to other amino acids for sows of mixed parity were according to NRC (2012) and kept similar across all dietary treatments. Sow BW and BF were measured during the start and end of each period. Number of piglets born live, still-born, mummies; number weaned; litter weights, and wean-to-estrus interval was recorded. Serum samples collected at the start and end of first gestation and lactation were analyzed for serum amino acids concentration.

Feeding low lysine diet in first phase (d0 – d35) of gestation did not affect (P = 0.227) the BW and BF changes in sow for that period. There was no effect (P> 0.05) of dietary treatments on sow body condition during both reproductive cycle. Total born, born alive, number weaned, number of mummies, stillborn and low viable piglets were not affected (P> 0.05) by dietary treatments. Litter weight gain and litter weight for each phases were not different (P> 0.05) among dietary treatments. Sows phase fed in the previous gestation and fed control diet in the previous lactation had 1 to 1.5 more (P < 0.05) piglets born alive in the subsequent parity than sows that were phase fed in the previous lactation period regardless of feeding regime in the previous gestation period. Principal component analysis of serum amino acid data revealed time-dependent difference (d0 vs d109 in gestation, d8 vs d18 in lactation). Also differences between gestation and lactation samples were also observed. However, the effects of phase feeding were not apparent.

Key words: Lysine, phase feeding, sow, litter, parity

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Introduction

Sows reproductive performance in terms of numbers of piglet weaned is steadily improving. The average number of piglets weaned per sow was 7.95 in 1990 (USDA, 1990) and for 2015 production year sows on national average weaned 10.3 pigs per litter (USDA, 2015). This improvement in performance is because the sows were selected for larger litter size and higher milk production. Better nutrition for the sows demand, housing and management strategies and reproductive technologies are key to those improvements. Modern sows are selected for leaner genetics and generally have larger body size and substantially low body fat and lower appetite. This is why it is challenging to manage and feed modern high prolific sows. Thus it is imperative to do continuous feeding and nutritional assessment to fulfill their demand (Kraeling and Webel, 2015).

Nutrient requirement of sow depends upon their age, stage of growth and production. It is important to formulate diets which are appropriate for each stages to fulfill these stage dependent requirements. Proper nutrition and management during gestation and lactation prepares sow for successful reproduction throughout their lifetime (Aherne, 2010). Currently the U.S. swine industry provides a constant level of nutritional and energy allowances to sows throughout the gestation period, with approximately 10 to 12 g/d of standardized ileal digestible (SID) lysine and 6,500 to 7,200 kcal/d of metabolizable energy (ME) in entire gestation period (Goodband et al., 2013). This leads to constant amino acid: energy ratio in the diet throughout the gestation period. When the requirement for energy increases by 25 to 35% from early to late gestation that of major limiting amino acids of gestating sows increases about two-fold from early to late gestation (Goodband et al., 2013). Nitrogen retention of whole body, including maternal and fetal protein pools, increases substantially from early gestation to late gestation. This has been documented as 0.84, 0.75, 1.00, and 1.36 g/d for day 10-40, 40-65, 65-90, and 90-114 of gestation, as expressed relative to day 65-90 (NRC, 2012). The fetal protein accretion was found to increase from 0.25 to 4.63 g/ day when we compare the requirements before and after day 69 of gestation (McPherson et al., 2004). It has also been shown that protein accretion rate in average individual mammary gland of pregnant gilts increased from 0.08 g/d before day 75 of gestation to 1.05 g/d after day 75 of gestation (Ji et al., 2006). The daily nitrogen gains in primiparous sows from all maternal and fetal tissues were 40 and 103 g/d respectively, before and after day 70 of gestation (Ji et al., 2005). This information points to the need to have higher protein levels in gestating sow diets only from the 69th day.

During late gestation and early lactation sow often becomes catabolic and mobilizes both protein and fat reserve to support fetal growth and milk production (Aherne and Williams, 1992., Pluske et al., 1998). Extended catabolic condition negatively affects longevity and productivity of sows (Foxcroft et al., 1995). Feeding a single gestation ration leads to overfeeding in early gestation and underfeeding in late gestation (Moehn et al., 2012). Feeding higher levels in early gestation could deleteriously affect embryo survival, especially in gilts (Heugten, 2000) as embryo survival decreased from 82.8% to 71.9% when feeding levels were increased from 1.5 kg to 3.0 kg daily (Dyck et al., 1980) as this also would accrue higher feed costs. Increasing

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lysine level and keeping energy intake constant has been shown to improve the birth weight of litters (Heo et al., 2008; Yang et al., 2008; Zhang et al., 2011)

Phase feeding of gestation sows is suggested to be a better management and nutritional practice to accommodate sow’s nutrient requirements and improve efficiency of sow productivity (review by Kim et al, 2013; Goodband et al., 2013; Moehn and Ball, 2013). However the sow production criteria has not been tested therefore a three phase sow feeding program; phase 1 (d 0- d35 of gestation), phase 2 (d 35- d 70) and phase 3 (d70-109) based on lysine level and balanced for other essential amino acid is expected to provide appropriate nutrition by closely matching nutrient need of the sow as well as the developing fetus.

As lactation progress, the milk production of sows also increases and peaks around third or fourth week of lactation which increases the demand for lysine (Noblet and Etienne, 1987). Modern sows wean more piglets than conventional sows with increase in the number of functional mammary glands (Kim et al., 2013). In this regard lactating sows will need additional nutrient supply not only to support the higher litter size but also for the growth of these mammary glands (Kim et al., 2013). It is reported that the mammary gland in sows continues to grow after farrowing with the increase in number of mammary epithelial in third and fourth week of lactation (Kim et al 1999). In order to meet the protein and lysine requirements of the lactating sow, Aherne, (2001) suggested that dietary protein (lysine) level should be 17.7 (0.97), 18.6 (1.00), 21.0 (1.12), and 21.4 (1.13) % in weeks 1, 2, 3, and 4 of lactation, respectively. To our knowledge, there is no reported information on phase feeding lactating sows. In order to maximize the amino acid intake during the peak lactation period when the demand is high, sows in this study were subjected to lactation feeding regimen; phase 1 (d 0- d6 of gestation), phase 2 (d 7- d 12) and phase 3 (d12-d 18) based on lysine level and balanced for other essential amino acid was studied.

Objectives

To evaluate the effects of phase feeding based on dietary lysine to multiparous gestating and lactating sows on sow and litter performance and piglet survivability.

Specific objectives:

a. To determine the effect of phase feeding dietary lysine to multiparous gestating and lactating sows on farrowing rate, sow body weight and backfat changes, wean-to-breed interval, litter size and weight and piglet survivability at birth and at weaning for the present and subsequent parity.

b. To determine the effect of phase feeding dietary lysine to multiparous gestating and lactating sows on plasma metabolomics during the period of gestation and lactation.

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Procedures

The experimental protocols used in this study were approved by the University of Minnesota Institutional Animal Care and Use Committee.

Animals and Management

The experiment was conducted in the swine research unit at the University of Minnesota’s Southern Research and Outreach Center in Waseca, Minnesota. A total of 240 mixed parity (1-7) sows (English Belle, GAP Genetics, Winnipeg, Manitoba, Canada) in six batches were used in the study. Sows were housed in individual stalls (0.61 m × 2.13 m) with an individual feeder and drinker from breeding until d 35 of pregnancy. Sows were fed once daily at 0730 h daily using automated trickle feeding system. Sows were moved to group pens (6.7 m × 12.8 m) on d 36, where they stayed until d 109 and were fed individually using electronic feed station (EFS, Osborne Industries, Inc., Osborne, Kansas). All sows were fed 1.81 kg from d 0 to d 56, 2.04 kg from d 56 to 84, 2.26 kg d 84 to d 96 and 2.5kg from d 96 to d 109 of gestation.

Sows were moved to farrowing crates (2.13 m long × 0.97 m high × 0.66 m wide) on d109 of gestation and were fed 2.27 kg of their assigned lactation diet starting on d 109 of gestation until farrowing. Post farrowing, the feed allowance was gradually increased to allow ad libitum feed intake of their lactation diet from day 4 until weaning. Sows were fed twice daily at 0800 h and 1500h. Piglets from the same dietary treatment groups were cross-fostered within 24 hours of birth and standardized to 12 pigs per sow. All piglets were processed (iron dextran injection, tail docking and navel disinfection) within 24 h of birth and surgical castrations performed on all male piglets around 1 week of age following the farm standard management practice. Heat lamps were provided around the time of farrowing till 48 h after birth. Piglets had access to heat pads in farrowing crate until weaning. Piglets with low birth weight (< 0.82 kg) were considered low viable (LV) piglets and were euthanized using CO2 chamber. Reasons for piglet mortality were recorded. The average lactation length in the farm was 19 days. After weaning sows were moved to individual crates and were checked daily for signs of estrus using mature boars. Sows were allowed free access to water throughout the study period.

All sows and piglets were monitored daily for general health and appropriate environment following sow gestation and lactation facility protocol approved in by University of Minnesota Institutional Animal Care and Use Committee.

Dietary treatment

Sows were blocked by parity, body weight, and backfat thickness at breeding and assigned one of the two dietary treatments during the gestation period in a randomized complete block design. Diets were formulated on standardized ileal digestible (SID) AA basis. The phase-fed sows (n = 121, BW = 227.0 ± 3.6 kg, BF = 16.5 ± 0.7 mm) received 0.4%, 0.57% and 0.7 % SID lysine diets in three different periods 1(d0 (breeding) - d35), 2 (d35- d70) and 3 (d70 – d109), respectively during gestation. The control group sows (n = 120, BW = 227.7 ± 3.5 kg, BF =16.3 ± 0.7 mm) were fed a 0.57% SID lysine diet throughout gestation. Sows were moved to farrowing crates on d109 and assigned to two different lactation feeding regimen in a 2 x 2 factorial arrangement (two level of gestation and two levels of lactation diet). Half of the sows

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from each gestation treatment group were assigned one of the two lactation treatment. During lactation period, control sows were fed a conventional lactation diet (1.0% standard ileal digestible (SID) lysine, 18% CP) throughout lactation or test diets (SID lysine 0.8%, 1.0%, and 1.2%) for phases 1(d 0-6), 2(d 7-12) and 3 (d 13 - weaning) respectively. All sows on were fed conventional control diets during successive gestation and lactation periods. Ratios of lysine to other amino acids for sows of mixed parity were according to NRC (2012) and kept similar across all dietary treatments.

Gestation Period Feed Regimen:

Control: single- phase feeding 0.57% standardized ileal digestible (SID) lysine

Treatment: three-phase feeding (0.4% SID lysine for day 0-35 of gestation, 0.57% SID lysine for day 35-70 of gestation; 0.7% SID lysine for day 70-109 of gestation)

Lactation Period Feed Regimen:

Control: single-phase feeding 1.0% SID lysine

Treatment: three-phase feeding (0.80% SID lysine for day 0-6 of lactation; 1.00% SID lysine for day 6-12 of lactation; 1.20% SID lysine for day 12-18 of lactation)

Data Collection

Feed sample analysis

Feed samples for each batch of all dietary treatments during gestation and lactation were collected; mixed and representative samples were used for analysis. Concentration of AA (AOAC 2006; method 982.30 E) and CP (AOAC 2006; method 990.03) in diet samples were analyzed at Experiment Station Chemical Laboratories (University of Missouri, Columbia, MO).

Sow and Litter Performance

All sows were weighed and backfat depth was determined at breeding and at the start and the end of all 3 phases of gestation and lactation. Backfat measurements were taken ultrasonically (Lean-Meater, Renco Corp., Minneapolis, MN) at the last rib about 6.5 cm from both side of the backbone using cooking oil as coupling fluid. The value from both left and right sides were averaged to obtain backfat depth. Feed allowed and wasted was also recorded on daily basis during lactation.

Litter weight was taken at birth and at the end of each phase until weaning. Total piglets born, born alive, mummies, stillborn, number and weight of the piglets cross fostered were also recorded. Incidence of death, probable cause and the weight of the dead piglets were also recorded. After weaning sows were monitored daily for post-weaning estrus and the dates were recorded to determine wean to estrus interval.

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Blood sampling and AA analysis

Blood samples were collected from 8 sows per treatment via jugular venipuncture 4 hour post feeding in BD vacutainer (Becton, Dickinson and Company, NJ) without anticoagulant at breeding and d 109 of gestation and at d 6 of lactation and at weaning. Blood samples were centrifuged at 1500 × g for 15 minutes and serum samples were stored at -80°C for later analysis. Serum samples were analyzed for concentration of free AAs and their percentage in total amino acid as described by Wang et al., (2016)

Concentrations of all free AA in serum were determined by liquid chromatography–mass spectrometry (LC-MS) using a modified method based on Márquez et al. (1986). Briefly, serum sample was prepared by mixing one volume of serum, one volume of 100 μM p-chlorol-L-phenylalanine (internal standard), and 18 volumes of 66% aqueous acetonitrile, and then centrifuging at 18,000 × g for 10 min to obtain the supernatant. Five µL of deproteinized sample was mixed with 40 μL of Na2CO3 (10 mM, pH = 11) and 100 μL of dansyl chloride (3 mg/mL in acetone). The mixture was incubated in a water bath at 60° C for 10 min followed by centrifugation at 18,000 × g for 10 min at 4° C. The supernatant was transferred to a HPLC vial and a 5 μL of aliquot was injected into an Acquity ultra-performance liquid chromatography (UPLC) system (Waters, Milford, MA) and separated in a BEH C18 column using a mobile phase gradient ranging from water to 95% aqueous acetonitrile containing 0.1% formic acid over a 10 min run. LC eluent was introduced into a Xevo-G2-S quadrupole time-of-flight mass spectrometer (QTOFMS, Waters) for accurate mass measurement and ion counting. Capillary voltage and cone voltage for electrospray ionization was maintained at 3 kV and 30 V for positive-mode detection. Source temperature and desolvation temperature were set at 120°C and 350°C, respectively. Nitrogen was used as both cone gas (50 L/h) and desolvation gas (600 L/h), and argon as collision gas. For accurate mass measurement, the mass spectrometer was calibrated with sodium formate solution (range m/z 50-1,000) and monitored by the intermittent injection of the lock mass leucine enkephalin ([M+H]+ = m/z 556.2771) in real time. Mass chromatograms and mass spectral data were acquired and processed by MassLynxTM software (Waters) in centroided format. Individual AA concentrations were determined by calculating the ratio between the peak area of AA and the peak area of internal standard and fitting with a standard curve using QuanLynxTM software (Waters).

Statistical Analysis

All the data were analyzed by SAS 9.4. Variables related to number of piglets, stillborn, mummy, and wean to estrus interval were considered as count data and analyzed by the GLIMMIXD procedure, and the remaining variables measured were considered as continuous variables and analyzed by the MIXED procedure. Number of piglets born alive, born in total, stillborn, and mummies for the current parity were analyzed as completely randomized block design with block (batch) as random effect and gestation treatment and parity as fixed effects with Poisson distribution. The remaining count data from the current parity and all count data in the subsequent parity were analyzed as split-plot design (main plot gestation and subplot lactation) with block (batch) as random effect and gestation, lactation, the interaction between gestation and lactation, and parity as fixed effects with Poisson distribution. Attempt

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was made to account for over dispersion by inclusion of the second random statement (random _residual_).

Sow body weight and backfat depth on days 35, 70 and 109 of gestation for the current parity were analyzed as completely randomized block design with repeated measures, whereas sow body weight, backfat depth, litter weight on days 6 and 12 and at weaning of lactation for the current parity, and sow feed intake during the 3 phases of lactation for both the current and subsequent parity were analyzed as split-plot design (main plot gestation and subplot lactation) with repeated measures. Sow body weight and backfat at breeding and their changes during gestation of the current parity were analyzed as completely randomized block design with block (batch) as random effect and gestation treatment and parity as fixed effects. The remaining continuous variables were analyzed as split-plot design (main plot gestation and subplot lactation) with block (batch) as random effect and gestation, lactation, the interaction between gestation and lactation, and parity as fixed effects. Means were separated by PDIFF option. A significant level was set at 0.05 and trend was considered if p-value between 0.05 and 0.1. Least square means and standard errors are presented

Principle component analysis of AA data was done by using SIMCA-P+TM software (version 11, Umetrics, Kinnelon, NJ) for all gestation and lactation samples.

Results and Discussion

 Sow and Litter Performance

Dietary effects on performance of sows and piglets during the first cycle of reproduction are shown in Tables 5 and 6. Feeding low lysine diet during the first phase (day 0 to 35 of gestation) did not affect (P = 0.227) the body weight and backfat change in sows for that period, which suggests that feeding low lysine diet in early gestation might not be detrimental to sows performance. Our data is in accordance with Moehn et al., (2012) who suggested a single phase feeding program leads to overfeeding in early gestation. Sows on phase feeding regimen gained more weight (Control vs. Phase feeding, 12.68 vs. 15.76 kg; P = 0.007) during the second phase (day 35 – 70) of gestation. Nevertheless, sows gained similar weight (Control vs. Phase feeding, 34.21 vs. 34.91 kg; P = 0.639) and backfat (Control vs. Phase feeding, 2.2 vs. 1.9 mm; P = 0.412) from breeding to day 109 of gestation. As gestation advanced sows gained more weight as indicated by the significant effect of time (P < 0.0001). It has been shown that the significant growth of the fetus and mammary gland takes place during late phase of gestation (McPherson et al., 2004, Ji et al., 2006).

Sow body weight and backfat depth were not different (P > 0.05) at all three phases of lactation. Sows that were phase fed in both gestation and lactation had the lower (P < 0.05) body weight loss than sows that were phase fed in gestation and fed control diet in lactation. Sows that were phase fed in both gestation and lactation lost least weight during the last phase (day 12- weaning) of lactation in comparison with other treatments, which emphasizes the increased requirement of nutrients in the peak lactation period (Aherne, 2001). Feed intake for all phases and total feed intake for the lactation period were not affected (P > 0.05) by dietary treatment. Sows that were phase fed in gestation and fed control diet took the shortest time to return to

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estrus than other groups and had significant lower (P < 0.05) weaning-to-estrus interval than sows that were phase fed during gestation and lactation.

Dietary treatment did not affect (P > 0.05) total born, number born alive, number weaned, number of mummies, stillborn and low viable piglets in this study. A study conducted in the Netherland (reported in National Hog Farmer, reviewed by Neutkens, 2006) used different levels of nitrogen (crude protein) and phosphorous in different phases of gestation and found no differences in number born alive, still born, number weaned and weaning weight. However the level of nutrients provided in late phases of gestation was not reported in this study. Litter weight and litter weight gain for each phases and overall litter weight gain were not affected (P > 0.05) by the dietary treatment.

Sow and litter performance in the subsequent reproductive cycle            Dietary effects on performance of sows and piglets during the subsequent reproductive cycle are shown in Table 7. Dietary treatments did not affect (P > 0.05) changes of sow body weight and back fat in gestation and at weaning and lactation sow feed intake during the 3 phases. However, sows that were phase fed in gestation and control fed in the previous cycle had higher (P < 0.05) overall feed intake than sows fed control diet in both gestation and lactation during the previous cycle.            Sows phase fed in the previous gestation and fed control diet in the previous lactation had 1 to 1.5 more (P < 0.05) piglets born alive in the subsequent parity than sows that were phase fed in the previous lactation period regardless of feeding regime in the previous gestation period. Number of piglets weaned, number of mummies and stillborn, litter weight at farrowing and weaning were not affected (P > 0.05) by dietary treatment imposed in the previous cycle.

Serum AA concentration

Figure 1 shows the principle component analysis (PCA) modeling of all gestation and lactation serum samples for free amino acid concentration used in the study. Principle component analysis modelling of serum AAs revealed time dependent difference (P > 0.05) in the concentration of AAs in gestation which is shown in Figure 2. Concentration of free glycine, alanine and ammonia were found to increase (P > 0.05) in serum during gestation whereas concentration of lysine, valine, leucine and isoleucine were found to decrease (P > 0.05) at d109 of gestation compared at breeding. However, the effect of dietary treatment was not observed in serum amino acid concentration for gestation serum samples. Similarly the time dependent difference (P > 0.05) in the concentration of free amino acids in was observed for lactation serum samples as shown in Fig3. Dietary treatment did not affect the concentration of serum free amino acids in lactation as shown in Fig4.

Summary

This current study highlights that feeding low protein (AA) diet in early gestation is not detrimental to sows performance. The effect of feeding high lysine diet in the later phases was not apparent in this study for sow body condition and reproductive performance. Phase feeding

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in gestation and control feeding in lactation showed improvements of 1 to 1.5 more piglets born alive in second cycle.

Optimizing feed efficiency by providing appropriate nutrient at appropriate time when it can be used effectively is important. Phase feeding sows during gestation and lactation is a nutritional management strategy in which pork producers can adjust the ingredients and chemical composition of the diets over time to fulfill the requirements of sows as well as the developing fetus. Most of the sow performance variable studied had clear parity effect which was predictable as sow body is dynamic. Sows used in this study ranged from parity 1-7 so phase feeding sows separated for different parity groups may be beneficial which should be weighed on with economic benefits and farm condition.

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Table 1. Ingredient and composition of (% as fed basis) experimental diets fed to gestating sows1

Ingredients Control Treatment DietPhase 1 Phase 2 Phase 3

Corn 65.03 69.05 65.03 60.62SBM2 10 6 10 14cDDGS3 20 20 20 20CWG4 1.5 1.5 1.5 1.5

Limestone 1.0 1.02 1.0 1.02Dicalcium Phosphate 1.2 1.25 1.2 1.25Lysine HCl 0.1 - 0.1 0.14DL-Methionine - - - 0.13L-Threonine - - - 0.12L-Tryptophan - - - 0.05Salt 0.35 0.35 0.35 0.35Premix5 0.5 0.5 0.5 0.5Tylan 0.125 0.125 0.125 0.125

CompositionME, kcal/kg 3,342 3,343 3,342 3,341Crude Protein, % 15.7 14.03 15.5 17.5Fermentable fiber, % 12.53 11.82 12.53 13.22Total Phosphorus 0.59 0.58 0.59 0.8STTD Phosphorus 0.35 0.35 0.35 0.53Calcium 0.75 0.75 0.75 0.64SID 6 , %

Crude Protein 12.49 11.09 12.49 13.86Lysine 0.57 0.7 0.57 0.7Methionine 0.25 0.23 0.25 0.4Threonine 0.44 0.38 0.44 0.61Tryptophan 0.12 0.10 0.12 0.19Arginine 0.73 0.62 0.73 0.85Histidine 0.36 0.32 0.36 0.40Isoleucine 0.50 0.43 0.50 0.57Leucine 1.41 1.31 1.41 1.50Methionine +Cysteine 0.47 0.44 0.47 0.63Phenylalanine 0.65 0.58 0.65 0.72

Phenylalanine +Tyrosine 1.12 1.00 1.12 1.24Valine 0.61 0.54 0.61 0.67

SID Lysine/ME 1.712 1.183 1.712 2.099 1Dietary treatments are as follows: Control and Phase 2 = 0.57% SID Lysine gestation diet; Phase 1= 0.4 % SID Lysine gestation diet; Phase 3 = 0.7 % SID Lysine gestation diet. 2SBM = 47.5% CP Soybean meal 3cDDGS = Corn dried distiller grains containing less than 4% oil , 4CWG = Choice white grease 5The vitamin and trace mineral premix provided the following (per kg of diet): vitamin A, 11,000 IU; vitamin D3, 2,756 IU; vitamin E,

55 IU; vitamin B12, 55µg; vitamin K, 4.4 mg riboflavin, 9.9 mg; pantothenic acid, 33 mg; niacin, 55 mg; choline, 495 mg; pyridoxine, 2.2 mg; folic acid, 1.65 mg; thiamine, 1.1 mg; biotin, 220 µg; I, 2.2 mg; Zn, 150 mg; Fe, 124 µg; Mn, 40 mg; Cu, 14.9 mg; and Se, 0.3 mg.

6SID = Standard Ileal Digestible value (NRC, 2012)

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Table2.Analyzed amino acid composition of experimental diets for gestation sows

Control TreatmentPhase 1 Phase 2 Phase 3

Crude Protein, % 15.43 13.41 15.43 15.51AA acids, %Taurine 0.16 0.15 0.16 0.10Hydroxyproline 0.02 0.03 0.02 0.07Aspartic acid 1.24 0.93 1.24 1.32Threonine 0.58 0.48 0.58 0.69Serine 0.69 0.58 0.69 0.70Glutamic acid 2.63 2.20 2.63 2.52Proline 1.13 1.02 1.13 1.05Lanthionine 0.00 0.00 0.00 0.00Glycine 0.61 0.50 0.61 0.65Alanine 1.00 0.89 1.00 0.93Cysteine 0.28 0.23 0.28 0.26Valine 0.74 0.61 0.74 0.75Methionine 0.26 0.22 0.26 0.33Isoleucine 0.61 0.49 0.61 0.60Leucine 1.68 1.48 1.68 1.55Tyrosine 0.54 0.46 0.54 0.45Phenylalanine 0.76 0.63 0.76 0.76Hydroxylysine 0.04 0.04 0.04 0.05Ornithine 0.01 0.01 0.01 0.01Lysine 0.70 0.51 0.70 0.82Histidine 0.41 0.34 0.41 0.46Arginine 0.81 0.61 0.81 0.85Tryptophan 0.17 0.13 0.17 0.20Total AA, % 15.07 12.54 15.07 15.121Dietary treatments2Percentage of Nitrogen (N × 6.25)Results are expressed on “as is” basis

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Table 3. Ingredient and composition of (% as fed basis) experimental diets fed to lactating sows1

Ingredients Control Treatment DietPhase 1 Phase 2 Phase 3

Corn 61.155 65.995 61.155 56.045SBM2 17.2 12.5 17.2 22.15cDDGS3 15 15 15 15

CWG4 3 3 3 3Limestone 0.88 0.88 0.88 0.88Dicalcium phosphate 1.15 1.20 1.15 1.10

Lysine HCl 0.46 0.35 0.46 0.56DL-Methionine 0.01 - 0.01 0.04L-Threonine 0.13 0.07 0.13 0.2L-Tryptophan 0.04 0.03 0.04 0.05Salt 0.35 0.35 0.35 0.35Premix5 0.5 0.5 0.5 0.5

Tylan 0.125 0.125 0.125 0.125C omposition ME, kcal/kg 3,381 3,382 3,381 3,379Crude Protein, % 18.01 16 18.01 20.12Fermentable fiber, % 13.07 12.24 13.07 13.94Total Phosphorus 0.66 0.65 0.66 0.67STTD Phosphorus 0.40 0.40 0.40 0.40Calcium 0.80 0.80 0.80 0.80SID 6 , %

Crude Protein 14.28 12.65 14.28 16.0Lysine 1.00 0.80 1.00 1.2Methionine 0.26 0.23 0.26 0.32Threonine 0.64 0.51 0.64 0.77Tryptophan 0.19 0.16 0.19 0.23Arginine 0.91 0.78 0.91 1.06Histidine 0.42 0.37 0.42 0.46Isoleucine 0.58 0.51 0.58 0.67Leucine 1.52 1.41 1.52 1.63Methionine+Cysteine 0.51 0.46 0.51 0.59Phenylalanine 0.77 0.69 0.77 0.86Phenylalanine+Tyrosine 1.27 1.14 1.27 1.42Valine 0.68 0.60 0.68 0.76

SID Lysine/ME 2.97 2.373 2.97 3.562Note: 1Dietary treatments are as follows: Control and Phase 2 = 18% CP, 1.0% SID Lysine lactation diet; Phase 1= 16% CP, 0.8 % SID

Lysine lactation diet; Phase 3 = 20 % CP, 1.2 % SID Lysine lactation diet.

2SBM = 47.5% CP Soybean meal 3cDDGS = Corn dried distiller grains containing less than 4% oil. 4CWG = Choice white grease. 5Premix supplied the following micronutrients (per kilogram of diet): 11,000 IU of vitamin A; 2,756 IU of vitamin D3; 55 IU of

vitamin E; 55µg of vitamin B12; 16,000 mg of riboflavin; 44.1 mg of pantothenic acid; 82.7 mg of niacin; Zn, 150 mg; 175 mg of Fe; 60 mg of Mn; 17.5 mg of Cu; 2 mg of I; and 0.3 mg of Se.

6SID = Standard Ileal Digestible value (NRC, 2012)

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Table 4. Analyzed amino acid composition of experimental diets for gestation sows

Control TreatmentPhase 1 Phase 2 Phase 3

Crude Protein, % 18.39 16.01 18.39 19.80AA acids, %Taurine 0.15 0.16 0.15 0.15Hydroxyproline 0.03 0.03 0.03 0.03Aspartic acid 1.63 1.22 1.63 1.77Threonine 0.80 0.64 0.80 0.89Serine 0.82 0.67 0.82 0.86Glutamic acid 3.07 2.50 3.07 3.26Proline 1.21 1.08 1.21 1.28Lanthionine 0.00 0.00 0.00 0.00Glycine 0.73 0.59 0.73 0.78Alanine 1.04 0.94 1.04 1.10Cysteine 0.30 0.26 0.30 0.32Valine 0.87 0.71 0.87 0.94Methionine 0.29 0.26 0.29 0.34Isoleucine 0.75 0.59 0.75 0.81Leucine 1.78 1.58 1.78 1.89Tyrosine 0.60 0.51 0.60 0.67Phenylalanine 0.89 0.73 0.89 0.96Hydroxylysine 0.04 0.05 0.04 0.04Ornithine 0.01 0.01 0.01 0.02Lysine 1.20 0.87 1.20 1.34Histidine 0.48 0.39 0.48 0.51Arginine 1.05 0.79 1.05 1.14Tryptophan 0.23 0.18 0.23 0.24Total AA, % 17.97 14.76 17.97 19.341Dietary treatments2Percentage of Nitrogen (N × 6.25)Results are expressed on “as is” basis

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Table 5. Effect of phase feeding in gestation on sow body condition and performance during the first cycleGestation P-Value

Control Phase Treatment (gestation)

Parity Time

Number of sows (N) 120 121Average parity 3.84 3.85

Sows body weight (BW), kgDay 0, breeding 227.7 ± 3.5 227.0 ± 3.6 0.7935 <0.0001Day 35, gestation 229.1 ± 2.7 227.0 ± 2.9

0.8440 <0.0001 <0.0001Day 70, gestation 241.8 ± 2.7 242.6 ± 2.9Day 109, gestation 262.2 ± 2.7 261.9 ± 2.9BW change (Day 0 - Day 35) 1.31 ± 1.45 0.25 ± 1.46 0.2270 <0.0001BW change (Day 35 - Day 70) 12.68 ± 1.40 15.76 ± 1.41 0.0074 0.4675BW change (Day 70 - Day 109) 20.15 ± 1.05 19.25 ± 1.07 0.4318 0.7298BW change (Day 0 – Day 109) 34.21 ± 2.11 34.91 ± 2.14 0.6390 <0.0001

Sows backfat (BF) depth, mmDay 0, breeding 16.3 ± 0.7 16.5 ± 0.7 0.6979 0.0393Day 35, gestation 15.7 ± 0.6 16.0 ± 0.6

0.7505 0.0831 <0.0001Day 70, gestation 16.7 ± 0.6 17.0 ± 0.6Day 109, gestation 18.6 ± 0.6 18.6 ± 0.6BF change (Day 0 - Day 35) 0.51 ± 0.36 -0.35 ± 0.34 0.5004 0.5648BF change (Day 35 - Day 70) 0.93 ± 0.53 0.98 ± 0.53 0.8619 0.5064BF change (Day 70 - Day 109) 1.63 ± 0.38 1.27 ± 0.39 0.3687 0.2283BF change (Day 0 – Day 109) 2.2 ± 0.53 1.86 ± 0.54 0.4122 0.1585

PigletsTotal born 13.59 ± 0.35 13.67 ± 0.37 0.8401 0.0989Number of born alive 11.31 ± 0.37 11.07 ± 0.38 0.4181 0.0001Mummy 0.73 ± 0.27 0.57 ± 0.21 0.2463 0.0040Stillborn 1.75 ± 0.21 2.14 ±0.26 0.1224 0.0092Low viable piglet2 0.96 ± 0.19 0.83 ± 0.17 0.3392 0.54171Dietary treatments = 1 Gestation; a, Control = conventional gestation sow diet based on 0.57% SID (standard ileal digestible lysine), b, phase = 3 phase feeding program; phase 1 (d0- d35 of gestation), phase 2 (d35- d 70) and phase 3 (d70-109) with 0.4%, 0.57% and 0.7% SID lysine respectively. Values are least square means ± standard error of mean2Piglets with body weight less than 0.82kg within 24 hours of birth were considered low viable piglets

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Table 6. Effect of phase feeding on sows reproductive performance during the first cycleTreatment1 P value

Gestation Control Control Phase PhaseGestatio

nLactatio

n

Gestation x

Lactation

Parity TimeLactation Control Phase Control Phase

Number of sows 57 63 60 61Average parity of sows 3.84 3.86 3.83 3.85PigletsNumber weaned 10.10 ± 0.36 10.13 ± 0.37 10.36 ± 0.38 9.98 ± 0.37 0.7973 0.3277 0.2515 0.0006Wean to estrus interval (day) 6.0 ± 0.49ab 5.6 ± 0.49ab 5.0 ± 0.47b 6.4 ± 0.55a 0.7647 0.2802 0.0342 0.4362

Litter weight (LW), kgLW Day 6 27.3 ± 1.55 27.9 ± 1.58 28.1 ± 1.50 27.7 ± 1.60

0.5921 0.9774 0.2984 <.0001LW Day 12 44.5 ± 1.55 45.3 ± 1.58 45.9 ± 1.50 44.9 ± 1.60LW Day 18 66.2 ± 1.55 67.6 ± 1.58 68.4 ± 1.50 66.8 ±1.60LW gain (Day 0 - Day 6) 11.2 ± 0.5 11.3 ± 0.5 11.5 ± 0.5 11.2 ± 0.5 0.4883 0.8293 0.2793 <.0001LW gain (Day 6 - Day 12) 17.3 ±0.7 17.8 ± 0.7 17.8 ± 0.7 17.5 ± 0.7 0.8316 0.9207 0.3841 0.0009LW gain (Day 12 - Day 18) 20.9 ± 1.0 21.5 ± 1.1 21.6 ± 1.1 21.1 ± 1.1 0.8481 0.9435 0.4656 0.0178LW gain (Day 0 – Day 18) 49.8 ± 1.7 50.5 ±1.8 51.2 ±1.8 49.8 ±1.8 0.5779 0.9714 0.2665 0.0002

Sows body weight (BW), kgDay 0, farrowing 247.3 ± 4.8 242.6 ± 4.9 246.5 ± 5.0 248.0 ± 5.0 0.3817 0.6579 0.5097 <.0001Day 6, lactation 255.3 ± 3.4 251.3 ± 3.5 252.6 ± 3.5 255.4 ± 3.6

0.9863 0.9889 0.1589 <.0001Day 12, lactation 255.8 ± 3.4 252.1 ± 3.5 251.7 ± 3.5 255.6 ± 3.6Day 18, lactation 253.3 ± 3.4 248.6 ± 3.5 247.8 ± 3.5 253.7 ± 3.6BW change (Day 0 - Day 6) 6.2 ± 1.7 4.4 ± 1.7 7.2 ± 1.7 5.5 ± 1.7 0.3966 0.1536 0.9743 <.0001BW change (Day 6 – Day 12) 1.1 ± 1.0 1.2 ± 1.1 0.1 ± 1.1 1.0 ± 1.1 0.5212 0.6219 0.6742 0.1143BW change (Day 12 - Day 18) -2.0 ± 0.9ab -3.0 ± 0.9ab -3.2 ± 0.9b -1.03 ± 0.9a 0.6410 0.4337 0.0411 0.1480BW change (Day 0 – Day 18) 4.3 ±2.5 1.5 ± 2.5 3.3 ± 2.6 5.1 ± 2.6 0.4871 0.7811 0.1517 <.0001

Sows backfat (BF) depth, mmDay 0, farrowing 17.5 ± 0.9 16.8 ± 0.9 16.9 ± 0.9 17.2 ± 0.8 0.9069 0.8258 0.4704 0.0928Day 6, lactation 17.4 ± 0.6 16.7 ±0.7 17.1 ± 0.7 17.8 ± 0.7

0.4992 0.9667 0.2084 <.0001Day 12, lactation 16.7 ± 0.6 16.1 ± 0.7 16.4 ± 0.7 17.05 ± 0.7

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Day 18, lactation 15.9 ± 0.6 15.2 ± 0.7 15.6 ± 0.7 16.2 ± 0.7BF change (Day 6 - Day 0) -0.4 ± 0.2 -0.6 ± 0.2 -0.2 ± 0.2 -0.3 ± 0.2 0.1560 0.1642 0.6089 0.0032BF change (Day 12 – Day 6) -0.7 ± 0.1 -0.6 ± 0.2 -0.7 ± 0.2 -0.7 ± 0.2 0.5375 0.8692 0.4832 0.0030BF change (Day 18 - Day 12) -0.9 ± 0.2 -0.8 ± 0.2 -0.7 ± 0.2 -0.8 ± 0.2 0.4880 0.6229 0.4194 0.0909BW change (Day 18 – Day 0) -2.0 ± 0.4 -2.2 ± 0.4 -1.6 ± 0.4 -1.9 ± 0.4 0.1828 0.2885 0.8608 <.0001

Lactation Feed intake (FI), kgFI (Day 0 – Day 6) 25.6 ± 1.70 26.2 ± 1.71 27.4 ± 1.73 26.7 ± 1.74

0.1899 0.6693 0.2422 <.0001FI (Day 6 – Day 12) 38.8 ± 1.70 38.4 ± 1.71 41.6 ± 1.73 39.6 ± 1.74FI (Day 12 – Day 18) 57.6 ± 1.70 59.7 ± 1.71 60.5 ± 1.73 58.2 ± 1.74FI (Day 0 – Day 18) 121.0 ± 4.1 124.5 ± 4.3 127.4 ± 4.3 122.3 ± 4.4 0.5270 0.8183 0.2070

1Dietary treatments = 1 Gestation; a, Control = conventional gestation sow diet based on 0.57% SID (standard ileal digestible lysine), b, phase = 3 phase feeding program; phase 1 (d0- d35 of gestation), phase 2 (d35- d70) and phase 3 (d70-109). 2, lactation; a, Control =conventional gestation sow diet based on 1.0 % SID (standard ileal digestible lysine) b, phase = 3 phase feeding program; phase 1 (d0- d6 of lactation), phase 2 (d6- d 12) and phase 3 (d12-weaning) with 0.8%, 1.0% and 1.2% SID lysine respectively. Values are least square means ± standard error of mean a,bMeans within a row with different superscripts differ (P < 0.05)

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Table 7. Effect of phase feeding on sows reproductive performance during the second cycleTreatment1 P value

Gestation Control Control Phase PhaseGestatio

nLactatio

n

Gestation x

Lactation

Parity TimeLactation Control Phase Control Phase

PigletsTotal born 13.76 ± 0.56 13.47 ± 0.53 14.76 ± 0.55 13.34 ± 0.52 0.3784 0.0596 0.2168 0.7126Number born alive 11.56 ± 0.49ab 11.26 ± 0.43b 12.55 ± 0.44a 11.02 ± 0.43a 0.4062 0.0186 0.1150 0.0038Mummy 0.57 ± 0.17 0.84 ± 0.17 0.76 ± 0.18 0.90 ± 0.18 0.4539 0.2248 0.6252 0.8910Stillborn 2.28 ± 0.37 2.12 ± 0.33 2.35 ± 0.35 2.29 ± 0.35 0.6851 0.7276 0.8438 0.0038Number weaned 10.15 ± 0.30 10.57 ± 0.28 10.56 ± 0.27 10.59 ± 0.29 0.4593 0.3319 0.3987 0.1071Litter weight at farrowing 16.5 ± 0.7 16.0 ± 0.6 17.5 ± 0.6 16.3 ± 0.6 0.2362 0.1178 0.5066 0.2185Litter weight at weaning 63.0 ± 2.2 65.3 ± 2.1 65.8 ± 2.0 66.7 ± 2.2 0.3672 0.3252 0.7530 <.0001

SowsBody weight (BW) at day109 256.7 ± 4.6 253.6 ± 4.6 249.0 ± 4.6 256.3 ± 5.0 0.5579 0.5049 0.1352 <.0001BW at weaning 248.1 ± 4.7 242.7 ± 4.3 240.9 ± 4.1 248.1 ± 4.4 0.8383 0.7661 0.0782 <.0001Backfat (BF) at d109 16.4 ± 1.0 15.5 ± 0.9 15.5 ± 0.8 16.3 ± 0.9 0.9444 0.9386 0.2334 0.5262BF at weaning 14.4 ± 0.9 13.1 ± 0.8 13.5 ± 0.8 14.3 ± 0.8 0.8066 0.6359 0.0870 0.0507BF change (D109 – weaning) -1.9 ± 0.5 -2.4 ± 0.4 -1.9 ± 0.4 -2.1 ± 0.4 0.6616 0.4391 0.6432 0.8718BW change (D109 - weaning) -8.4 ± 3.8 -10.0 ± 4.5 -7.8 ± 3.5 -7.8 ± 3.5 0.6058 0.7060 0.6935 0.0032

Lactation Feed intake (FI), kgFI (Day 0 – Day 6) 23.4 ± 2.65 25.3 ± 2.53 25.2 ± 2.49 25.6 ± 2.57

0.1775 0.9682 0.1196 <.0001FI (Day 6 – Day 12) 36.4 ± 2.65 37.8 ± 2.53 39.0 ± 2.49 36.7 ± 2.57FI (Day 12 – Day 18) 49.7 ± 2.65 52.2 ± 2.53 55.8 ± 2.49 52.1 ± 2.57FI (Day 0 – Day 18) 105.3 ± 9.9a 115.0 ± 9.7ab 119.2 ± 9.6b 110.6 ± 9.8ab 0.3435 0.8828 0.0159

1Dietary treatments = 1 Gestation; a, Control = conventional gestation sow diet based on 0.57% SID (standard ileal digestible lysine), b, phase = 3 phase feeding program; phase 1 (d0- d35 of gestation), phase 2 (d35- d70) and phase 3 (d70-109). 2, lactation; a, Control =conventional gestation sow diet based on 1.0 % SID (standard ileal digestible lysine) b, phase = 3 phase feeding program; phase 1 (d0- d6 of lactation), phase 2 (d7- d 12) and phase 3 (d12-weaning) with 0.8%, 1.0% and 1.2% SID lysine respectively.Values are least square means ± standard error of mean a,bMeans within a row with different superscripts differ (P < 0.05).

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Fig1. Principle component analysis (PCA) modeling of all gestation and lactation serum samples for free amino acid concentration D0 = day of breeding, d109= day 109 of gestation, d 18= day of weaning and d6 = a 6 of lactation.

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Fig2. Principle component analysis (PCA) modeling of gestation serum samples for free amino acid concentration Control_d0 = day 0 (breeding) start of the trial, Control_d109 = sows on control diet on day 109 of gestation, Phase_d0 = day 0 (breeding) start of the trial, phase_d109 = sows that were phase fed on day 109 of gestation

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Fig3. Principle component analysis (PCA) modeling of lactation serum samples on d 6 and d 18 for free amino acid concentration

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Fig4. Principle component analysis (PCA) modeling of lactation serum samples on for free amino acid concentration.CTL-CTL = Sows fed control diet in gestation and lactationCTL-PHF = Sows fed control diet in gestation and phase diets in lactationPHF-CTL = Sows fed phase diets in gestation and control diet in lactationPHF-PHF = Sows fed phase diets in gestation and lactation

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