Effects of dietary yeast mannan-rich fraction supplementation on growth performance, intestinal morphology, and lymphoid tissue characteristics in weaned piglets challenged with Escherichia Coli F4

doi:10.21203/rs.3.rs-3994291/v1

Download PDF

Research Article

Effects of dietary yeast mannan-rich fraction supplementation on growth performance, intestinal morphology, and lymphoid tissue characteristics in weaned piglets challenged with Escherichia Coli F4

https://doi.org/10.21203/rs.3.rs-3994291/v1

This work is licensed under a CC BY 4.0 License

Journal Publication

published 29 May, 2024

Read the published version in Tropical Animal Health and Production →

You are reading this latest preprint version

We evaluated the effects of supplementing yeast mannan-reach-fraction on growth performance, jejunal morphology and lymphoid tissue characteristics in weaned piglets challenged with E. Coli F4. A total of 20 crossbred piglets were used. At weaning, piglets were assigned at random to one of four groups: piglets challenged and fed the basal diet supplemented with yeast mannan-rich fraction (C-MRF, n = 5); piglets challenged and fed the basal diet (C-BD, n = 5); piglets not challenged and fed the basal diet supplemented with yeast mannan-rich fraction (NC-MRF, n = 5), and piglets not challenged and fed the basal diet (NC-BD). Each dietary treatment had five replicates. On days 4, 5 and 10, piglets were orally challenged with 10⁸ CFU/mL of E. Coli F4. C-MRF piglets had higher BW (p = 0.002; interactive effect) than C-BD piglets. C-MRF piglets had higher (p = 0.02; interactive effect) ADG in comparison with C-BD piglets. C-MRF piglets had higher (p = 0.04; interactive effect) ADFI than C-BD piglets. The diameter of lymphoid follicles was larger (p = 0.010; interactive effect) in the tonsils of C-MRF piglets than C-BD piglets. Lymphoid cells proliferation was greater in the mesenteric lymphnodes and ileum (p = 0.04 and p = 0.03, respectively) of C-MRF piglets. A reduction (p > 0.05) in E. Coli adherence in the ileum of piglets fed MRF was observed. In conclusion, the results of the present study demonstrate that dietary yeast mannan-rich fraction supplementation was effective in protecting weaned piglets against E. Coli F4 challenge.

Mannan oligosaccharide

growth

immunity

Enterotoxigenic Escherichia coli

weanling pig

Weaning is one of the most challenging phases in pig production (Lallès et al. 2007). During this time, piglets are subjected to numerous stressors factors, e.g., abrupt separation from the sow, transportation, co-mingling with unacquainted piglets, exposure to new pathogens and food-associated antigens, and change from the highly digestible sow’s milk to a dry solid diet, composed with less digestible proteins (Campbell et al. 2013). This invariably triggers an immune response and disrupts the structure and function of the intestine (Liao 2021; Tang et al. 2022). Further, weaning has been associated with a state of microbial dysbiosis (Chen et al. 2017), which in association with disrupted epithelial barrier function allows pathogens to cross the intestinal epithelium resulting in inflammation, mal-absorption, post-weaning diarrhea and growth check (Pluske et al. 2018). With respect to pathogens, enterotoxigenic Escherichia Coli F4 (E.coli F4; ETEC) is the most diffuse etiological agents associated with post-weaning diarrhea in pigs (Luise et al. 2019), causing an economic burden for pig farmers (Fairbrother et al. 2005).

Antibiotics have long been included in weaned piglet diets, either prophylactically or sub-therapeutically, to prevent ETEC infection and promote growth performance (Verstegen and Williams 2002). However, the increasing concern regarding antimicrobial resistance has led to reduced use or ban of in-feed antibiotics in some countries (Guevarra et al. 2018). The use of prebiotics has attracted considerable attention as a potential alternative for the use of in-feed antibiotics. In this regard, Mannan oligosaccharides (MOS) are prebiotics that derive from the cell wall of the yeast Saccharomyces cerevisiae and serve as nutritional sources for beneficial microorganisms (e.g., Lactobacilli and Bifidobacteria) in the gastrointestinal tract (Halas and Nochta 2012) MOS can improve intestinal health by changing intestinal microbiota composition and inhibiting the adherence to enterocytes of gram-negative pathogens that express type-1 fimbriae (Borowsky et al. 2009; Zha et al. 2023). There is a wealth of evidence demonstrating that weaned piglets fed MOS-supplemented diet have improved body-weight gain, feed conversion and health (Spring et al. 2015). Likewise, MOS has been shown to improve intestinal mucosal barrier function in piglets by regulating intestinal microbiota balance and immune responses (Duan et al. 2019).

In recent years, research efforts have led to the isolation of unique carbohydrates in the outer cell walls of a specific strain of Saccharomyces cerevisiae, giving rise to the second generation of MOS, yeast-derived mannan rich fraction (MRF) (Spring et al. 2015). MRF supplies highly soluble mannans, allowing the partial exposition of β-glucans. This results in enhanced prebiotic effect and local immune-modulatory actions. The use MRF have been shown improved intestinal morphology and permeability, and reduced intestinal inflammatory response in weaned piglets (Song et al. 2019). Further, supplying MRF to piglets altered cecal microbial community structure and improved jejunal morphology early post-weaning (Fouhse et al. 2019). Supplementation with MRF effectively boosted the immune responses and improved nutrient utilization in weaned piglets challenged with PRRSV (Che et al. 2012). However, there is a lack of studies evaluating the effects of dietary MRF supplementation of weaned piglets challenged with ETEC.

Based on the foregoing, the objective of this study was to evaluate the effects dietary MRF supplementation on growth performance, jejunal morphology, and lymphoid tissue characteristics (lymphoid cell proliferation and lymphoid follicle diameter) in weaned piglets challenged with E. Coli F4.

Ethics approval

All animal experimental design and procedures were approved by the Ethics Committee on Animal use of the Federal University of Lavras under the protocol n° 012/17.

Animals, housing, and diets

The experiment was conducted at the nursery facility of the Swine Experimental Center at the Department of Animal Science, Federal University of Lavras (21°14ꞌ43ꞌꞌS44°59ꞌ59ꞌꞌW, Lavras, Minas Gerais, Brazil). A total of 20 crossbred barrows (Duroc × Landrace × large white) weighing in average 7.50 ± 0.66 kg and weaned at approximately 24 days were used. The piglets were housed individually in pens (1.1 m × 1.5 m) with slatted plastic flooring, self-feeders (5-feeding spaces), and drinking nipple. The room temperature was controlled manually by opening or closing the windows. Temperature thermometers were used to indicate when the windows should be closed or opened. A corn–soybean meal basal diet was formulated to meet or exceed the nutritional requirements of nursery piglets, according to the Brazilian Poultry and Swine Tables (Rostagno et al. 2017; Table 1). Two different diets were used during the experimental period: pre-starter I (0-4days) and pre-starter II (4–11 days). Piglets had ad libitum access to feed and water throughout the experiment. No antibiotics were included in the basal diet. There was inclusion of pharmacological doses of zinc oxide in the basal diet (2000 and 1500 ppm of zinc in pre-starter I and pre-starter II diets, respectively). As a prophylactic measure against respiratory diseases, all piglets were administered intramuscularly 0.15 ml tulathromycin (Draxxin®, 100 mg/ml of tulathromycin, Zoetis®, São Paulo, Brazil) at weaning.

Table 1

Composition of the experimental diets.
Ingredients (%)	Pre-starter I	Pre-starter II
Corn 7,88% CP	42.3	56.9
Soybean meal 45% CP	15.7	19.7
Micronized soybean	20.3	9.0
Spray-dried plasma	5.0	3.0
Dried milk	9.0	6.0
Soybean oil	-	1.42
Vitamin premix¹	0.05	0.05
Micromineral premix²	0.1	0.1
Dicalcium phosphate 18.5% P	1.0	1.2
Limestone	1.0	0.9
Salt	0.4	0.5
Zinc oxide 80%	0.3	0.2
L-Lysine-HCL	0.29	0.47
DL-methionine	0.15	0.17
L-treonine	0.06	0.12
L-tryptophan	-	0.01
L-valine	-	0.02
Inert	0.2.	0.2
Total	100	100
Calculated levels (%)
ME (kcal/kg)	3490.2	3400.0
Fat	6.6	5.9
Crude protein	23.4	19.8
SID Lysine	1.65	1.48
SID Methionine	0.43	0.43
SID Met + Cys	0.82	0.74
SID Threonine	0.88	0.79
SID Tryptophan	0.25	0.22
SID Arginine	1.41	1.16
SID Valine	1.02	0.86
SID Isoleucine	0.78	0.73
SID Leucine	1.77	1.54
SID Histidine	0.59	0.59
SID Phenyalanine	1.01	0.85
Crude fibre	1.76	2.07
Lactose	10.00	5.00
Total Calcium	0.85	0.80
Available Phosphorus	0.45	0.40
Sodium	0.40	0.35
Copper (mg/kg)	6.08	5.50
Zinc (mg/kg)	2024.8	1529.0
¹The vitamin premix provided the following quantities of vitamins per kg of complete diet: 30.000.000 IU vitamin A, 6.000.000 IU vitamin D3, 180.00 IU vitamin E, 6.700 mg vitamin K3, 5.250 mg, 11.88 g vitamin B2, 47.5 g pantothenic acid, 6.880 vitamin B6, 68.750 mcg vitamin B12, 75 g nicotinic acid, 6.880 mg folic acid, 810 mg biotin. ²The micro mineral premix provided the following quantities of micro minerals per kg of complete diet: 80 g iron, 15 g copper, 41.3 g manganese, 91.2 g zinc, 330 g cobalt, 1.209 mg iodine, 351 mg selenium.

Experimental design and treatments

The experiment design was as randomized complete block, with blocks based on initial body weight, in a 2 x 2 factorial arrangement of treatments. The first factor was dietary MRF (Non-supplemented basal diet or basal diet supplemented with 0.1% MRF). The second factor was ETEC challenge (unchallenged or challenged with E. coli F4). The four experimental groups were as follows: piglets challenged and fed the basal diet supplemented with 0.1% MRF (HyperGen®, Biorigin, São Paulo, Brazil) (C-MRF, n = 5); piglets challenged and fed the basal diet without inclusion of MOS (C-BD, n = 5); piglets not challenged and fed the basal diet supplemented with 0.1% MRF (NC-MRF, n = 5), piglets not challenged and fed the basal diet without supplementation (NC-BD). Each dietary treatment had five replicates. On days 4, 5 and 10, piglets were inoculated by oral gavage with 10⁸ CFU/mL of E. Coli F4. The enterotoxogenic E. Coli strain (F4+, LT+, STa + and STb+) was isolated by the Laboratório de Sanidade Suína (FMVZ/USP). The enterotoxigenic E. coli was confirmed by PCR genotyping as genes expressing F4 fimbrial antigen. Prior to oral inoculation, E. coli F4 was grown overnight in MacConkey agar medium at 37°C using 0.3 mL of inoculum from stock. Fallowing centrifugation, the supernatant was discarded and the pellet obtained. The cells were washed three times with 30 mL of sterilized cold phosphate buffer solution (PBS), and after serial dilution a suspension containing 10⁸ CFU/mL of E. coli F4 was used.

Growth performance

Feed intake and individual body weight were recorded on days 0, 4 (before challenge), and 11. Likewise, the feed provided and refusals were evaluated daily. Thus, the average daily gain (ADG), the average daily feed intake (ADFI) and feed conversion ratio (FCR) were calculated.

Sample collection

On day 11 of the trial, all piglets were humanely euthanized by electrical stunning and exsanguination. A midline abdominal incision was made and the gastrointestinal tract collected. Thereafter, approximately 2-cm segment of the mid-jejunum were dissected, rinsed with 0.9% cold phosphate buffer solution (PBS) and fixed in 10% formaldehyde solution for morphology measurements. Samples of ileum, tonsils, mesenteric lymph nodes and spleen were collected and stored in 10% buffered formaldehyde. Additionally, samples of cecal digesta were collected and immediately stored at -80°C for microbiota and volatile fatty acids analyses.

Histology analyses

Samples of jejunal tissue were fixed in 10% formaldehyde solution for 48h and transferred to 70% alcohol solution until the slides were prepared. Sections (4 µm) were obtained and stained with hematoxylin and eosin, as previously described (Pan et al. 2016). The slides were photographed using trinocular microscope (CX31, Olympus Optical do Brasil Ltda., São Paulo, SP, Brazil) coupled with a high-definition camera (SC30, Olympus Optical do Brasil Ltda., São Paulo, SP, Brazil). The heights of 10 well-orientated villi and their adjoining crypts were measured using the AxionVision SE64 4.9.1 software. Villus height was defined as the distance between the crypt mouth and villi tip. Crypt depth was defined as the infolding between two villi. The ratio of villus height to crypt depth was also determined. Measurements of lymphoid follicles in the ileum, tonsils, mesenteric lymphnodes, and spleen samples were conducted using Image-Pro® Express software. The vertical (D1) and horizontal (D2) dimensions of the follicles were determined by measuring the greatest visual distances between their endpoints. Three lymphoid follicles were selected at random for measurement. When there were less than three lymphoid follicles in a tissue sample, all follicles were measured.

Immunohistochemistry

The anti-proliferating cell nuclear antigen immunohistochemistry technique was used to evaluate the proliferation of lymphoid cells in spleen, mesenteric lymph node, tonsils and ileum samples. Briefly, histological sections were adhered to silanized slides, dewaxed in xylene, rehydrated in graded alcohol followed by distilled water. Thereafter, the slides were placed in citrate buffer (pH 6.0) and subjected to microwave

for antigen recovery. To block endogenous peroxidase activity, slides were incubated with 3% hydrogen peroxide solution for 30 min at room temperature. After washing in phosphate buffer saline (PBS, 0.01 M pH 7.2), the sections were incubated with Mouse Recombinant Monoclonar anti-PCNA (Agilent Dako, Santa Clara, CA, USA). The slides with primary antibody were incubated overnight) at 4ºC. Subsequently, the sections were incubated with the secondary antibody (Envision, Agilent, Santa Clara, CA, USA), for 1 h. For detection of E. coli in jejunal samples a specific polyclonal antibody (DAKO B0357, Agilent Dako, Santa Clara, CA, USA) was used. 3,3′-diaminobenzidine (DAB) was used as substrate to visualize the bound antibodies. The computer program Image-Pro Express (version 6.0 for Windows, Media Cybernetics, Bethesda, MD, USA) was used to detect the average integrated optical density of the positive products. For each organ, images were obtained of three random lymphoid follicles and three different image fields for each follicle. The cells marked by the anti-PCNA antibody were counted.

Short-chain fatty acids concentration

The analysis of short-chain fatty acids (i.e., acetic, propionic and butyric) was performed on samples of cecal digesta. For this purpose, 4 mL of formic acid (17%) was added to 2 g of cecal digesta in order to extract and preserve fatty acids. Centrifugation was carried out at 2500 rpm and the supernatants stored at -20°C until gas chromatography analysis (Agilent 7890A GC®, Santa Clara, CA, USA).

Statistical analyses

All residues were tested for normality and homogeneity using the Shapiro-Wilk test. The data were analyzed using the SAS software statistical package 9.3 (SAS, Cary, NC). The data was analyzed as a randomized complete block design using the PROC GLIMMIX procedure in SAS 9.3 (SAS Inst. Inc., Cary, NC). The piglet was considered as the experimental unit. All data were described as LSMEANS. The means were compared by Tukey test. Differences were considered significant if p < 0.05, with a trend toward significance at 0.05 ≤ p ≤ 0.10.

Growth performance

The effects of MRF on growth performance are shown in Table 2. From days 0–4, no statistical difference was observed for any performance variable. In the overall experimental period (0–11 days), there was an interaction between dietary MRF and ETEC challenge, with C-MRF piglets having higher (p = 0.002) final BW than C-BD piglets. Likewise, there was a significant interaction between dietary MRF and ETEC-challenge in ADG of piglets; C-MRF piglets had higher (p = 0.02) ADG in comparison with the C-BD group. The FCR tended to be improved (p < 0.08) in piglets from the C-MRF group compared to the C-BD group. There was a MRF x ETEC-challenge interaction in ADFI, with C-MRF piglets having higher (p = 0.04) ADFI when compared to C-BD group.

Table 2

Effect of dietary MRF supplementation on growth performance in weaned piglets challenged with ETEC
Treatments							P values
Itens	C-BD	NC-BD	C-MRF	NC-MRF	SEM	MRF	C	MRF x C
Initial BW, kg	7.56	7.57	7.63	7.46	0.18	0.950	0.940	0.940
0–4 days
BW, kg	7.88	8.07	8.28	7.99	0.01	0.243	0.777	0.108
ADG, kg	0.07	0.12	0.16	0.13	0.13	0.142	0.472	0.243
ADFI, kg	0.17	0.22	0.19	0.21	0.5	0.941	0.308	0.584
FCR	1.80	1.82	1.19	1.78	0.01	0.386	0.521	0.440
0–11 days
BW, kg	9.47 ^b	10.32 ^ab	10.96 ^a	10.38 ^ab	0.66	0.007	0.002	0.002
ADG, kg	1.97 ^b	2.82 ^ab	3.47 ^a	2.88 ^ab	0.43	0.017	0.347	0.020
DFI, kg	0.17 ^b	0.25 ^ab	0.31 ^a	0.26 ^ab	0.03	0.017	0.350	0.020
FCR	1.59	1.63	1.13	1.53	0.08	0.017	0.032	0.082
Abbreviations: BW, body weight; ADG, average daily gain; ADFI, average daily feed intake; FCR, feed conversion ratio; C-BD, piglets challenged with E. Coli F4 and fed the basal diet; NC-BD, piglets not challenged and fed the basal diet; C-MRF, piglets challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; NC-MRF, piglets not challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; SEM, standard error of the mean.
^a,bMeans within rows with different superscript are different at p ≤ 0.05.

Jejunal morphology and lymphoid follicles diameter

The results of jejunal morphology are presented in Table 3. No interactive effect between diet and ETEC challenge was observed. Villus height tended to be longer (p = 0.08) in piglets supplemented with MRF challenged or not with ETEC. All other jejunal measurements were not different (p > 0.05) among treatments. The results of lymphoid follicles diameter are shown in Table 4. The diameter of lymphoid follicles was larger (p = 0.010; interactive effect; Fig. 1) in the tonsils of piglets from the C-MRF group when compared to C-BD group. A tendency for MRF x ETEC-challenge interaction was observed for lymphoid follicles diameter in the mesenteric lymphnodes, with C-MRF piglets having larger (p = 0.71) lymphoid follicles than C-BD piglets.

Table 3

Effect of dietary MRF on jejunal morphology
Treatments							p values
Item	C-BD	NC-BD	C-MRF	NC-MRF	SEM	MRF	C	MRF x C
VH, µm	342.5	429.8	469.2	446.7	23.5	0.088	0.645	0.256
CD, µm	324.0	330.3	341.0	334.6	24.0	0.481	0.658	0.733
VH:CD	1.0	1.3	1.4	1.4	0.07	0.177	0.752	0.433
Abbreviations: VH, villus height; CD, crypt depth, VD:CD, villus height to crypt depth ratio; C-BD, piglets challenged with E. Coli F4 and fed the basal diet; NC-BD, piglets not challenged and fed the basal diet; C-MRF, piglets challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; NC-MRF, piglets not challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; SEM, standard error of the mean.

Table 4

Effects of supplementing MRF on lymphoyd follicles diameter in weaned piglets challenged with E. Coli F4
Treatments							P-value
Item	C-BD	NC-BD	C-MRF	NC-MRF	SEM	MRF	C	MRF x C
Tonsils	286.56 ^b	375.00 ^a	442.33 ^a	359.22 ^a	22.47	0.030	0.928	0.010
Mesenteric lymphnodes	318.55	301.00	318.93	393.30	17.88	0.069	0.248	0.071
Ileum	1076.42	834.19	1087.32	1171.68	10.06	0.217	0.839	0.197
Spleen	218.20	202.39	234.22	250.16	18.45	0.125	0.959	0.421
Abbreviation: C-BD, piglets challenged with E. Coli F4 and fed the basal diet; NC-BD, piglets not challenged and fed the basal diet; C-MRF, piglets challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; NC-MRF, piglets not challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; SEM, standard error of the mean.
^a,bMeans within rows with different superscript are different at p ≤ 0.05

Lymphoid cell proliferation and E. coli adherence

The results of lymphoid cell proliferation are presented in Table 5. A diet x ETEC-challenge interaction was found for lymphoid cells proliferation in the mesenteric lymphnodes and ileum (p = 0.04 and p = 0.03, respectively), indicating that challenged-piglets supplemented with MRF had higher number of lymphoid cell proliferation than challenged-piglets fed the basal diet. A treatment effect of diet was observed for lymphoid cell proliferation, with piglets supplemented MRF having greater number of proliferating lymphoid cells in spleen, mesenteric lymphnods and tonsils. The immunohistochemistry analysis showed a reduction in E. Coli adherence in the ileum of piglets fed MRF (Fig. 2).

Table 5

Effects of supplementing MRF on lymphoid follicles proliferation in weaned piglets challenged with *E. Coli F4*
Treatments							p values
Variável	C-BD	NC-BD	C-MRF	NC-MRF	SEM	MRF	C	MRF x C
Spleen	305.83	137.43	335.41	360.38	32.00	0.032	0.216	0.190
Ileum	565.28 ^a,b	275.72 ^b	637.16 ^a	709.70 ^a	36.10	0.001	0.111	0.042
Mesenteric Lynphnode	315.54 ^a,b	167.19 ^b	407.76 ^a,b	519.03 ^a	42.60	0.001	0.678	0.033
Tonsil	439.23	426.16	489.98	645.10	24.06	0.008	0.135	0.168
Abbreviation: NC-BD, piglets not challenged and fed the basal diet; C-MRF, piglets challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; NC-MRF, piglets not challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; SEM, standard error of the mean.
^a,b,cMeans within rows with different superscript are different at p ≤ 0.05

Short-chain fatty acids concentration

The results of cecal volatile fatty acids concentration are exhibited in Table 6. Overall, dietary MRF supplementation had no effect on the concentration of volatile fatty acids. The ETEC challenge increased total concentration of volatile fatty acids (p < 0.05).

Table 6

Effects of supplementing MRF on cecal concentrations of short-chain fatty acids in weaned pigs challenged with *E. Coli F4*
Treatments							p values
Item	C-BD	NC-BD	C-MRF	NC-MRF	SEM	MRF	C	MRF x C
Acetate	1047.85	430.78	978.14	426.31	61.902	0.906	0.002	0.928
Propionate	610.44	201.55	688.02	250.71	72.618	0.550	0.001	0.893
Butyrate	268.62	127.99	270.42	106.22	63.309	0.841	0.006	0.803
Total	1926.92	760.32	1793.93	783.25	62.072	0.927	0.002	0.865
Abbreviation: C-BD, piglets challenged with E. Coli F4 and fed the basal diet; NC-BD, piglets not challenged and fed the basal diet; C-MRF, piglets challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; NC-MRF, piglets not challenged and fed the basal diet supplemented with 0.1% mannan-rich fraction; SEM, standard error of the mean.

At weaning piglets are exposed to a host of biological, social and environmental challenges (Campbell et al. 2013). Further, during this stressful phase piglets are susceptible to various bacterial pathogens; among these, ETEC is an important causative agent of diarrhea and decreased growth performance in newly weaned piglets (Fairbrother et al. 2005). Our results demonstrated that supplementation of weaned piglets with MRF is a potential nutritional strategy to mitigate the physiological stress arising from weaning. Supplementing MRF to weaned piglets challenged with ETEC resulted in higher ADG and BW compared to piglets challenged and not supplemented with MRF. Further, piglets fed MRF and challenged with ETEC consumed more feed compared to piglets challenged and not supplemented. Our results disagree with(Song et al. 2019) who observed that MRF supplemented diet had no effects on growth performance of unchallenged piglets. Likewise, it has been shown that supplementing piglets with MRF had no effect on overall growth performance of unchallenged piglets (Fouhse et al. 2019). Conversely, dietary MOS significantly improved gain to feed ratio of weaned piglets challenged with Salmonella Typhimurium (Burkey et al. 2004). These conflicting results may be due to differences in mannan oligosaccharide sources, the process involved in its productiom, piglet age, additive amount, and health status. With respect to the latter, it has been reported that dietary MOS supplementation are more effective in improving performance when piglets are raised under challenging conditions (Zhao et al. 2012). Moreover, the effectiveness of MOS as growth promoter is considered to be greater during early stages of the nursery period (Zhao et al. 2012).

The maintenance of adequate intestinal morphology is of paramount importance for nutrient digestion and absorption of weaned piglets (Tang et al. 2022). Weaning stress affects intestinal health, which is represented by functional changes in the small intestine, including villous atrophy and increased crypt depth (Campbell et al. 2013). Villus height directly affects surface area and is linearly associated with nutrient absorption (Pluske et al. 2018). Therefore, the reduction in villus height and villus height to crypt depth ratio compromise intestinal mucosal function, resulting in reduced intestinal absorption capacity (Kim et al. 2012). In the present study no interactive effect between MRF and ETEC challenge was found for jejunal morphology. However, supplementation with MRF tended to increase villus height in jejunum of both challenged and unchallenged piglets. Of note, (Fouhse et al. 2019) reported that jejunal villus height was significant longer in piglets fed diets supplemented with MRF, but the supplementation had no effect on crypt depth and villus height to crypt depth ratio. The lack of significant effect on villus height in our study might be due the low number of piglets used. More studies with larger number of animals are needed to evaluate the effects of dietary supplementation with MRF on jejunal morphology of challenged piglets.

There is mounting evidence indicating that dietary MRF can enhance immune system activity in animals (Che et al. 2012; Spring et al. 2015). Dietary MOS can have both an indirect and a direct effect on the immune response of pigs (Halas and Nochta 2012). Of particular interest, it has been shown that MOS can increase the proliferation of isolated lymphocytes (Davis et al. 2004). Corroborating this notion, in our study the diameter of lymphoid follicles was greater in tonsils of challenged piglets fed MRF. Further, we observed a higher proliferation of lymphoid cells in mesenteric and ileum lymphnodes of challenged piglets fed MRF. Taking together these results indicate an enhanced immune response in piglets supplemented MRF.

MOS is considered to have potential antimicrobial properties (Borowsky et al. 2009). Due to the β-glucan content in MOS it can effectively bind type-1 fimbriae on pathogenic bacteria preventing their attachment to the intestinal epithelium (Halas and Nochta 2012). MRF is more purified than MOS, providing therefore a great source of attachment for pathogenic bacteria expressing type-1 fimbriae (e.g., E. Coli F4) (Spring et al. 2015). In broiler chickens, dietary supplementation of MRF reduced Campylobacter colonization (Corrigan et al. 2017). Likewise, Salmonella prevalence and shedding was decreased in pigs supplemented with MRF (Andrés-Barranco et al. 2015). Dietary MRF supplementation altered cecal microbial community structure in weaned piglets (Fouhse, 2019). Corroborating the aforementioned effects, we observed that piglets supplemented with MRF had reduced ETEC adherence in the ileum.

Dietary MOS supplementation can promote the growth of beneficial bacteria (e.g., Lactobacillus and Bifidobacterium), which by MOS fermentation produces short-chain fatty acids, i.e., acetate, propionate and butyrate (Halas and Nochta 2012; Zhuang et al. 2020). Reported studies demonstrate that short-chain fatty acids play important roles in the maintenance of intestinal health by providing energy to enterocytes and gut microflora (Canfora et al. 2015). In our study, dietary MRF supplementation had no effect on cecal short-chain fatty acids concentration. Surprisingly, there was an effect of ETEC-challenge on short-chain fatty acids production. It is worth mentioning that in our study it was not possible to determine the effect of MRF on Lactobacillus counts due to an exacerbated growth of Lactobacillus colonies in the culture plate. Therefore, the mechanism underpinning these results is yet to be determined.

Our study strengthened the notion that dietary MRF supplementation is a promising nutritional strategy to replace the use of in-feed antibiotics and alleviate the biological stressor arising from weaning. Notwithstanding this, the results of the present study should be seen in light of some potential limitations. The main limitation was the low number of animals evaluated, which may have precluded that some of the results differed statistically. Further, we were not able to evaluate whether MRF supplementation enhanced the proportion of Lactobacillus in the cecum. More studies using larger number of animals and evaluating the growth of beneficial bacteria are warranted to further confirm the beneficial effects of dietary MRF supplementation for challenged piglets.

The results of the present study demonstrate that dietary MRF is effective in mitigating the biological stress arising from weaning. Supplementing MRF protected weaned piglets from E. Coli F4 challenge. Challenged piglets fed MRF had increased growth performance, and compared to challenged piglets not supplemented MRF. In order to strengthen the present results, additional studies using a larger number of animals are warranted.

Funding

The authors acknowledge the National Council for Scientific and Technological Development (CNPq) and Coordination for the Improvement of Higher Education Personnel (CAPES) for the financial support.

Author contribution

D.F.L., V.S.C.., D.L.R.: conceptualization, data curation, analysis, investigation, writing, review and editing

V.S.C., D.L.R: supervision, conceptualization, resources, and methodology

V.S.C., D.L.R., D.F.L: data curation, writing.

P.C.B., K.U.U., M.S.V., S.R.S.J., M.R., J.A.B.: review, investigation, writing-original draft preparation

All authors reviewed, edited, and approved the final document.

Data availability statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Conflict of interest

The authors declare no conflict of interest.

Andrés-Barranco S, Vico JP, Grilló MJ, Mainar-Jaime RC (2015) Reduction of subclinical Salmonella infection in fattening pigs after dietary supplementation with a ß-galactomannan oligosaccharide. Journal of Applied Microbiology 118:284–294. https://doi.org/10.1111/jam.12713
Borowsky L, Corção G, Cardoso M (2009) Mannanoligosaccharide agglutination by Salmonella enterica strains isolated from carrier pigs. Brazilian Journal of Microbiology 40:458–464. https://doi.org/10.1590/s1517-83822009000300007
Burkey TE, Dritz SS, Nietfeld JC, et al (2004) Effect of dietary mannanoligosaccharide and sodium chlorate on the growth performance, acute-phase response, and bacterial shedding of weaned pigs challenged with Salmonella entericaserotype Typhimurium1. Journal of Animal Science 82:397–404. https://doi.org/10.1093/ansci/82.2.397
Campbell JM, Crenshaw JD, Polo J (2013) The biological stress of early weaned piglets. J Anim Sci Biotechnol 4:2–5. https://doi.org/10.1186/2049-1891-4-19
Canfora EE, Jocken JW, Blaak EE (2015) Short-chain fatty acids in control of body weight and insulin sensitivity. Nature Reviews Endocrinology 11:577–591. https://doi.org/10.1038/nrendo.2015.128
Che TM, Song M, Liu Y, et al (2012) Mannan oligosaccharide increases serum concentrations of antibodies and inflammatory mediators in weanling pigs experimentally infected with porcine reproductive and respiratory syndrome virus. Journal of Animal Science 90:2784–2793. https://doi.org/10.2527/jas.2011-4518
Chen L, Xu Y, Chen X, et al (2017) The Maturing Development of Gut Microbiota in Commercial Piglets during the Weaning Transition. Frontiers in Microbiology 8:1–13. https://doi.org/10.3389/fmicb.2017.01688
Corrigan A, Corcionivoschi N, Murphy RA (2017) Effect of yeast mannan-rich fractions on reducing Campylobacter colonization in broiler chickens. Journal of Applied Poultry Research 26:350–357. https://doi.org/10.3382/japr/pfx002
Davis ME, Maxwell C V., Erf GF, et al (2004) Dietary supplementation with phosphorylated mannans improves growth response and modulates immune function of weanling pigs. Journal of Animal Science 82:1882–1891. https://doi.org/10.2527/2004.8261882x
Duan X, Tian G, Chen D, et al (2019) Mannan oligosaccharide supplementation in diets of sow and (or) their offspring improved immunity and regulated intestinal bacteria in piglet. Journal of Animal Science 97:4548–4556. https://doi.org/10.1093/jas/skz318
Fairbrother JM, Nadeau É, Gyles CL (2005) Escherichia coli in postweaning diarrhea in pigs: an update on bacterial types, pathogenesis, and prevention strategies . Animal Health Research Reviews 6:17–39. https://doi.org/10.1079/ahr2005105
Fouhse JM, Dawson K, Graugnard D, et al (2019) Dietary supplementation of weaned piglets with a yeast-derived mannan-rich fraction modulates cecal microbial profiles, jejunal morphology and gene expression. Animal 13:1591–1598. https://doi.org/10.1017/S1751731118003361
Guevarra RB, Hong SH, Cho JH, et al (2018) The dynamics of the piglet gut microbiome during the weaning transition in association with health and nutrition. Journal of Animal Science and Biotechnology 9:54. https://doi.org/10.1186/s40104-018-0269-6
Halas V, Nochta I (2012) Mannan oligosaccharides in nursery pig nutrition and their potential mode of action. Animals 2:261–274. https://doi.org/10.3390/ani2020261
Kim JC, Hansen CF, Mullan BP, Pluske JR (2012) Nutrition and pathology of weaner pigs: Nutritional strategies to support barrier function in the gastrointestinal tract. Anim Feed Sci Technol 173:3–16. https://doi.org/10.1016/j.anifeedsci.2011.12.022
Lallès J-P, Bosi P, Smidt H, Stokes CR (2007) Weaning — A challenge to gut physiologists. Livestock Science 108:82–93. https://doi.org/10.1016/j.livsci.2007.01.091
Liao SF (2021) Invited review: Maintain or improve piglet gut health around weanling: The fundamental effects of dietary amino acids. Animals 11:1–16. https://doi.org/10.3390/ani11041110
Luise D, Lauridsen C, Bosi P, Trevisi P (2019) Methodology and application of Escherichia coli F4 and F18 encoding infection models in post-weaning pigs. Journal of Animal Science and Biotechnology 10:1–20. https://doi.org/10.1186/s40104-019-0352-7
Pan L, Ma XK, Wang HL, et al (2016) Enzymatic feather meal as an alternative animal protein source in diets for nursery pigs. Animal Feed Science and Technology 212:112–121. https://doi.org/10.1016/j.anifeedsci.2015.12.014
Pluske JR, Turpin DL, Kim J-C (2018) Gastrointestinal tract (gut) health in the young pig. Animal Nutrition 4:187–196. https://doi.org/10.1016/j.aninu.2017.12.004
Rostagno HS, Albino LFT, Donzele JL, et al (2017) Tabelas Brasileiras Para Aves e Suínos: Composição de Alimentos e Exigências Nutricionais, 4rd edn. UFV, Viçosa
Song M, Fan Y, Su H, et al (2019) Effects of Actigen, a second-generation mannan rich fraction, in antibiotics-free diets on growth performance, intestinal barrier functions and inflammation in weaned piglets. Livestock Science 229:4–12. https://doi.org/10.1016/j.livsci.2019.09.006
Spring P, Wenk C, Connolly A, Kiers A (2015) A review of 733 published trials on Bio-Mos®, a mannan oligosaccharide, and Actigen®, a second generation mannose rich fraction, on farm and companion animals. Journal of Applied Animal Nutrition 3:1–11. https://doi.org/10.1017/jan.2015.6
Tang X, Xiong K, Fang R, Li M (2022) Weaning stress and intestinal health of piglets: A review. Frontiers in Immunology 13:1–14. https://doi.org/10.3389/fimmu.2022.1042778
Verstegen MWA, Williams BA (2002) Alternatives to the use of antibiotics as growth promoters for monogastric animals. Animal Biotechnology 13:113–127. https://doi.org/10.1081/ABIO-120005774
Zha A, Tu R, Qi M, et al (2023) Mannan oligosaccharides selenium ameliorates intestinal mucosal barrier, and regulate intestinal microbiota to prevent Enterotoxigenic Escherichia coli -induced diarrhea in weaned piglets. Ecotoxicology and Environmental Safety 264:115448. https://doi.org/10.1016/j.ecoenv.2023.115448
Zhao PY, Jung JH, Kim IH (2012) Effect of mannan oligosaccharides and fructan on growth performance, nutrient digestibility, blood profile, and diarrhea score in weanling pigs. Journal of Animal Science 90:833–839. https://doi.org/10.2527/jas.2011-3921
Zhuang Z, Ding R, Peng L, et al (2020) Genome-wide association analyses identify known and novel loci for teat number in Duroc pigs using single-locus and multi-locus models. BMC Genomics 21:1DUMM. https://doi.org/10.1186/s12864-020-6742-6

Download PDF

Journal Publication

published 29 May, 2024

Read the published version in Tropical Animal Health and Production →

Editorial decision: Major revision with re-assessment
24 Mar, 2024
Reviewers agreed at journal
07 Mar, 2024
Reviewers invited by journal
05 Mar, 2024
Editor assigned by journal
29 Feb, 2024
First submitted to journal
27 Feb, 2024

You are reading this latest preprint version

Effects of dietary yeast mannan-rich fraction supplementation on growth performance, intestinal morphology, and lymphoid tissue characteristics in weaned piglets challenged with Escherichia Coli F4

Status:

Journal Publication

Version 1

Abstract

Figures

Introduction

Materials and methods

Results

Discussion

Declarations

References

Status:

Journal Publication

Version 1