Enterococcus faecium supplementation in sows during gestation and lactation improves the performance of sucking piglets

Abstract The aim of this study was to evaluate the effects of Enterococcus faecium DSM 7,134 supplementation on the performance of sows and their litters. A total of 15 primiparous sows (Landrace × Yorkshire) were randomly divided into three treatments with five replicates. Dietary treatments were: CON, basal diet; E1, CON + 0.025% E. faecium; E2, CON + 0.05% E. faecium. No significant differences were observed on body weight and feed intake of lactating sows with E. faecium supplementation, but linearly increased the sow apparent total tract digestibility (ATTD) of dry matter (DM), nitrogen (N) and gross energy (GE; p < .05), and decreased piglets pre‐weaning mortality (p < .05). Piglets from E. faecium‐supplemented sows linearly increased weaning weight, average daily gain (ADG) and gain:feed ratio (p < .05), as well as linearly decreased diarrhoea score (p < .05) in the first weaning week. Piglets from E. faecium‐supplemented sows linearly increased faecal Lactobacillus and Enterococci counts (p < .05), while linearly decreased faecal Escherichia coli counts (p < .05) after weaning. In conclusion, dietary supplementation of E. faecium improved the ATTD of DM, N and GE in lactating sows, as well as improved body weight, ADG and shifted faecal microbiota in their litters.

sows' diet (Scharek et al., 2005). Probiotics supplementation in sows' diet seems to be a useful way in establishing beneficial bacterial species and reducing pathogen load in piglets. However, studies on the effect of E. faecium supplementation in sows are relatively few.
The body condition of lactation sows is heavily depended on feed intake and nutrient utilization, due to high mobilization during lactation (Nelssen, 1999). Improving feed intake and/or nutrient digestibility may have beneficial effects on sows' performance. Dietary E. faecium supplementation had beneficial effects on feed intake and weight performance of primiparous sows (Böhmer, Kramer, & Roth-Maier, 2006), as well as nutrient digestibility of weaning and growing pigs (Yan & Kim, 2013;Zhang et al., 2014). Lactation sows have severe catabolic conditions, due to massive milk production with limited nutrient intake (Kim and Easter, 2003). If feed intake or/and feed digestibility efficiency are limiting factors leading to catabolic condition, improving nutrient utilization is vital to sow. If the digestibility of nutrient in the conventional sow diet can be improved by dietary E. faecium supplementation, then total gross energy (GE) available to sows will enhance without increasing feed intake. In addition, the E. faecium also can transfer to piglets by contact with maternal faeces (Jadamus, Vahjen, & Simon, 2001), the indirect colonization via sow faeces may influence piglet performance and health status (Taras, Vahjen, Macha, & Simon, 2006). Therefore, the objective of this study was to evaluate the effects of E. faecium supplementation on performance and nutrient digestibility of sows, as well as growth performance and health status of piglets.

| MATERIAL S AND ME THODS
The experimental protocol used in this study was approved by the Animal Care and Use Committee of Dankook University.

| Source of Enterococcus Faecium
The E. faecium DSM 7,134 used in this study was provided by a commercial company (Schaumann Agri International GmbH, Pinneberg, Germany), which is composed of spray-dried spore-forming with at least 1.0 × 10 10 cfu/g of live E. faecium contained.

| Experimental design, animals and housing
A total of 15 sows (Landrace × Yorkshire) were randomly divided into three treatments with five replications. The treatment diets were fed 14 days before farrowing until weaning (weaning at 21 day).
Diets were formulated (Tables 1 and 2) to meet or exceed the nutrient requirements of pigs (NRC, 2012). Sows were feed on a commercial gestation and lactation feed (Table 1) in mash form. During gestation, sows were housed individually in stalls of 2.20 × 0.60 m 2 . The stall had partly slatted floors that consisted of a 0.84 m concrete solid floor and a 1.25 m concrete slatted floor. Approximately 10 day before parturition, sows were moved to farrowing crates, each with 2.20 × 1.80 m 2 . Temperature in the farrowing room was maintained at a minimum of 20°C. Feeds in 1 ml of PBS were serially diluted from 10 -1 to 10 -7 , and plated on bile esculin azide agar plates in duplicates for 48 hr at 37°C. No E. faecium counts were detected in the CON diet. The E. faecium counts were 2.72 × 10 8 and 2.75 × 10 8 cfu/kg in the E1 gestation and lactation diet, and 5.40 × 10 8 and 5.35 × 10 8 cfu/kg in the E2 gestation and lactation diet.

| Chemical analysis, sampling and measurements
Gross energy was determined by measuring the heat of combustion in the samples using a bomb calorimeter (Parr 6100; Parr instrument Co. Creep feed was not given to piglets during the lactation period, and sow milk was the only feed available during lactation. From day 21 to 35, faecal score of weaning pigs was recorded three times per day by the same person, according to the method described by Huang et al. (2015), the scores were as follows: 1 = well-formed faeces (hard or soft, formed, and moist stool that retains its shape), 2 = sloppy faeces (unformed stool that assumes the shape of the container) and 3 = diarrhoea (liquid stool that can be poured).
To determine the apparent total tract digestibility (ATTD) of dry matter (DM), nitrogen (N) and gross energy (GE), chromium oxide was added to the diets at 2 g/kg, as an indigestible marker (Fenton & Fenton, 1979). Sows were fed the diets for 7-day, followed by faecal grab sampling via rectal massage. All feed and faecal samples were stored at −20°C until analysis. Before chemical analysis, faecal samples were thawed and dried at 70°C for 72 hr, after which they were finely ground to a size that could pass through a 1-mm screen.
Chromium was analysed by UV absorption spectrophotometry (UV-1201; Shimadzu, Tokyo, Japan) following the method described by Williams, David, and Iismaa (1962). The digestibility was calculated according to the following formula: where analysis was immediately carried out according to the method described by Böhmer et al. (2006). One gram of faecal samples was diluted with 9 ml of 1% peptone broth to the dilution step from 10 -1 to 10 -7 .

| Statistical analysis
Both sow and piglet performance data were analysed with SAS 2003 (v. 9.1, SAS Institute Inc.) using the mixed GLM procedure. Sow BW and backfat data were analysed using a repeated measurement method. The method included diet as a fixed effect whereas sow and period were included as random effects. The sows were used as the experiment unit.

| Growth performance and nutrient digestibility in lactating sows
Dietary E. faecium supplementation had no significant differences in BW, BW loss, feed intake or backfat thickness loss of sows (Table 3).
Linear and quadratic effects were observed in pre-weaning mortality (p < .05) of piglets with the increasing level of E. faecium supplementation, and linear effects were observed in ATTD of dry matter, nitrogen and energy (p < .05; Table 4).

| Growth performance and faecal score in piglets
On weaning day, BW and ADG of piglets were higher in sows receiving the E. faecium supplemented diets compared with the CON diet (Table 5). Similarly, during day 22 to 35, linear trend were observed in ADG (p < .10) and linear effects were observed in G:F (p < .05) with E. faecium supplementation. During day 22 to 28, diarrhoea score linearly decreased (p < .05) with increasing levels of E. faecium supplementation.

| Faecal microbiota and blood immune parameters in lactating sows and piglets
In lactating sows, a quadratic effect was observed in Enterococci counts (p < .05), while no differences were observed in faecal Lactobacillus or E. coli counts among treatments (Figure 1a-c). On weaning day, E. faecium suplementation linearly increased fecal No differences were observed in blood immune parameters of sows and piglets with E. faecium supplementation (Table 6).

TA B L E 3
Effects of Enterococcus faecium supplementation on performance in sows (Taras et al., 2006). Probiotics can transfer to piglets by contact with maternal faeces (Jadamus et al., 2001), which may be accompanied by the beneficial effects on faecal microbiota, immunogenic factors and diarrhoea incidence (Schanler, 2000).

The increased faecal
Lactobacillus and Enterococci counts, and decreased E. coli counts of piglets on weaning day may explain the decreased pre-weaning mortality with dietary E. faecium supplementation. During lactation, body loss in sows is mainly due to high milk yield and relatively low feed intake (Lallès et al., 2007), the adequate feed intake for lactation sows is crucial to guarantee their performance. Alexopoulos et al. (2004) reported that there was an increase in feed intake and a decrease in weight loss in lactating sow with B. licheniformis and B.
subtilis blend supplementation. Other studies also confirmed the decreased weight loss of sow with probiotics supplementation during lactation (Kreuzer & Zerhusen, 1995). However, no significant differences were observed in weight loss or feed intake with E. faecium supplementation in this study. The different results may be due to different probiotics strain used, dose level and diet composition.
In this study, BW and ADG of weaning pigs were linearly increased with the increasing level of E. faecium in the diet of lactating sow.
Similar results were also reported by Alexopoulos et al. (2004), who indicated that sow administrated with Bacillus and E. faecium had lower pre-weaning mortality and higher weaning weight in piglets. Taras et al. (2006) reported the administration of E. faecium to sows and their piglets led to decreased piglet mortality and reduced pre-and post-weaning diarrhoea. Baker et al. (2013) also reported that sows with Bacillus supplementation improved litter weaning weight, ADG, and decreased mortality. The improved BW and ADG of piglets in this study may be due to improved nutrient digestibility of sows, and finally lead to better milk production. We know that there is a strong relationship between piglets' BW gain and milk production and constituent (Noblet, Dourmad, & Etienne, 1990). However, milk production and constituent were not measured here, which is the limitation of this study.
Dietary E. faecium supplementation linearly increased the ATTD of DM, N and GE in lactating sows, which was consistent with Zhang et al. (2014), who reported that the ATTD of N and GE was enhanced in weaning pigs with E. faecium supplementation. Yan and Kim (2013) also reported that dietary E. faecium supplementation increased the ATTD of DM, N and GE in growing pig. E. faecium is a normal microorganism in swine intestine, which produce lactic acid to reduce intestinal pH and inhibit the load of invasive pathogens (Canibe & Jensen, 2003), thus may be a reason to explain the improving nutrient digestibility in this study.

TA B L E 6 Effects of Enterococcus faecium supplementation to sows on blood immune parameters in sows and piglets
The gastrointestinal and lymphoid systems are the largest immunologically competent organs, the development and composition of the gastrointestinal microbiota are the principal factors influencing maturation and optimal development of immunologically systems (Cho & Kim, 2014). In this study, E. faecium supplementation showed only minor changes in the gut of sows with a slight reduction in E. coli counts and a slight increase in Lactobacillus counts. The gastrointestinal flora of adult sows has stabilized, unlike piglet, a fundamental change by using probiotics is unlikely (Gedek, 1993), which was confirmed by Sarabia, Villar, Magboo, and Roxas (1997).
The piglet gut is sterile in utero and becomes colonized after birth mainly by bacteria acquired from the sow and sow faeces (Mackie et al., 1999). The early development of the gastrointestinal microbiota and colonization by environmental bacteria have long-term effects on the host and immune development of the neonate (Tannock, 2005;Thompson, Wang, & Holmes, 2008), as well as regulates host metabolism, growth and susceptibility to disease (Konstantinov et al., 2006;Marques et al., 2010;Turnbaugh et al., 2006). Previous studies reported the transfer of Bacillus from sow to piglet via the faecal-oral route (Baker, Davis, Spencer, Moser, & Rehberger, 2008 (Broom, Miller, Kerr, & Knapp, 2006;Scharek, Guth, Filter, & Schmidt, 2007). In this study, no differences were observed in serum IgG, IgA, IgM or lymphocyte concentration with E. faecium supplementation. However, Szabó et al. (2009)

| CON CLUS ION
Our studies suggested that the supplementation of E. faecium in the diet of gestation and lactating sows had no significant effects on BW, BW loss, ADFI and backfat thickness of sows, but decreased pre-weaning mortality, improved BW, ADG and shifted faecal microbiota in piglets, as well as improved the digestibility of DM, N and GE in lactating sow.

ACK N OWLED G EM ENTS
We thank our lab members' kind help during the experiment.

CO N FLI C T O F I NTE R E S T
All authors have no potential conflict of interest to statement.

E TH I C A L S TATEM ENT
The authors confirm that the ethical policies of the journal, as noted on the journal's author guidelines page, have been adhered to and the appropriate Ethical Review Committee approval has been received. The Korean National Research Council's guidelines for the Care and Use of Laboratory Animals were followed.