
==== Front
Poult Sci
Poult Sci
Poultry Science
0032-5791
1525-3171
Elsevier

S0032-5791(24)00774-0
10.1016/j.psj.2024.104195
104195
MICROBIOLOGY AND FOOD SAFETY
Beneficial effects of duck-derived lactic acid bacteria on growth performance and meat quality through modulation of gut histomorphology and intestinal microflora in Muscovy ducks
Li Zhaolong 497377512@qq.com
*1
Zhou Haiou *
Liu Wenjin †
Wu Huini *
Li Cuiting *
Lin Fengqiang *
Yan Lu *
Huang Chenyu *
⁎ Institute of Animal Husbandry and Veterinary Medicine of Fujian Academy of Agricultural Sciences, Fujian, Fuzhou 350013, China
† Center for Animal Disease Control and Prevention of Changji Hui Autonomous Prefecture, Xinjiang, Changji 09942339853, China
1 Corresponding author: 497377512@qq.com
13 8 2024
11 2024
13 8 2024
103 11 10419529 5 2024
5 8 2024
© 2024 The Authors
2024
https://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Duck-derived lactic acid bacteria (DDL) are a crucial beneficial bacterium in the intestines, contributing significantly to the health of ducks. However, the mechanism by which these DDL improves the growth performance and meat quality of Muscovy duck is not clear. In this study, A total of 800 male Muscovy ducks, initially weighing 50.15 ± 5.37 g, were randomly allocated into 4 groups, each with 4 replicates, consisting of 50 ducks per replicate. The control group consumed deep well water, while the experimental groups were given water supplemented with 1%, 3%, and 5% DDL (1.59×108 CFU/mL). The study duration was 70 d. The results revealed that Muscovy ducks drinks with the DDL significant reduced the feed conversion ratio (FCR) (P < 0.05) and increased the sweetness and richness of duck meat, among which the 5% drinking group has the most significant difference. Further study finding, the DDL significantly increased the height of villi, the ratio of villi height/crypt depth (V/C) on jejunum and colon, and the ratio of acidic mucus, neutral mucus, and glycogen to tissue area in both the duodenum and ileum of Muscovy ducks, and significantly decreased the tunel positive cells. Moreover, DDL significantly enhanced the abundance of genus beneficial bacterium (Bacillus, lentilactobacillus, Bacterodies, Lactobacillus) on duodenum and ileum. Additionally, drink with the DDL elevated the level of IgG in blood and the immune indices of the thymus and the fabricius bursa (P<0.05). Meanwhile, the meat composition analysis demonstrated that Muscovy duck drinks with the DDL raised the level of the saturated fatty acid rate(C12:0), and polyunsaturated fatty acid (C18:2 n-6 and C20:5 n-3,), and the monounsaturated (C18:1 n-7, and C18:1 n-9). Furthermore, correlation analysis finding that the growth performance of Muscovy ducks was positively correlated with the height of villi, the ratio of villi height/crypt depth (V/C), the abundance of genus beneficial bacterium. And the meat quality of Muscovy ducks has positively correlated with genus beneficial bacterium in intestinal, glutamic acid, saturated fatty acid rate and polyunsaturated fatty acid. This finding suggest DDL is an effective strategy to improve the growth performance and meat quality of Muscovy ducks by gut histomorphology and intestinal microflora.

Key words

duck-derived lactic acid bacteria
Muscovy duck
growth performance
meat quality
==== Body
pmcINTRODUCTION

Lactic acid bacteria (LAB), is a type of nonspore-forming, Gram-positive bacteria that ferment sugars to produce lactic acid, which is widely distributed in nature and encompasses a rich diversity of species, with over 200 species divided into 18 genera (Filippis et al., 2020). LAB plays an indispensable and vital role in the physiological functions of livestock and poultry (Meruvu and Harsa., 2023). They are commonly found in the gastrointestinal tract of livestock and poultry, producing a significant impact on their overall health (Mani-López et al., 2022).

Therefore, LAB is recognized as one of the earliest feeds additives in China (Li et al., 2023).It produces active substances like lactic acid, hydrogen peroxide, and bacteriocin, which can inhibit harmful bacteria, reduces oxygen levels in the gut, lower pH levels, creates a favorable environment for beneficial bacteria, and supports their growth (Zhang et al., 2023). LAB is commonly used as a substitute for antibiotics and play a vital role in the ecological breeding of livestock and poultry. Moreover, they can regulate the balance of the normal flora system in the gut, maintaining a healthy microecological environment in animals' bodies, which contributes to the improved gastrointestinal function, food digestion, and overall health enhancement (Silpa et al., 2022). Previous studies have shown that adding LAB to chicken feed can enhance feed intake and daily weight gain (Reuben et al., 2019). Likewise, supplementation of duck feed with lactic acid bacteria has been found to increase beneficial bacteria in duck intestines, resulting in improved slaughter rates and feed efficiency (Xu et al., 2024). Recent research has further revealed that the inclusion of intestinal beneficial microorganisms, such as lactic acid bacteria, in Muscovy duck diets through fermentation can improve growth performance and reduce feed-to-meat ratios (Li et al., 2024).

LAB not only positively impacts the growth performance of Muscovy ducks, but also play a role in regulating fat metabolism in livestock and poultry to enhance meat quality (Xu et al., 2023). For example, they can influence the intramuscular fat content by affecting fat synthase activity and the expression of genes related to lipid synthesis. Probiotics, such as LAB, are also involved in metabolizing and absorbing macromolecules like host fats, leading to a reduction in saturated fatty acids and an increase in polyunsaturated fatty acids (Heir et al., 2022). When probiotics are added to chicken feed, there is an increase in total fatty acids in chicken meat, with a higher ratio of unsaturated to saturated fatty acids. Drinking of LAB results in upregulation of genes like THRSP and PPARγ for unsaturated fatty acids, while downregulating genes like FAS for saturated fatty acids, thus influencing fat metabolism and lipid metabolism (Johnson et al., 2023). Moreover, LAB aid in fat oxidation into smaller molecules, enhancing meat flavor and quality (S and R, 2022). Previous studies have shown that feeding Lactobacillus Johnson to broilers boosts the levels of taste substances like inosine, proline, and phenylalanine, improving meat flavor (Patterson and Burkholder, 2003). Additionally, Lactobacillus produces acetic acid, butyric acid, and propionic acid in the intestinal tract of livestock and poultry, which can be broken down into smaller molecular substances like aldehydes, ketone, alcohols, and hydrocarbons, significantly contributing to meat flavor formation (Ferrer-Lorente et al., 2007).

While the positive effects of drink with the DDL on the growth performance of Muscovy ducks and meat quality of livestock and poultry have been demonstrated, the specific impacts on Muscovy ducks are still not fully understood. This study aimed to investigate the benefits of incorporating Muscovy duck Lactobacillus into their drinking water, assessing performance indicators and meat quality. Through examining changes in gastrointestinal structure, intestinal microbiota, and analyzing meat composition, the study offers a preliminary analysis of the mechanisms underlying the influence of DDL drinking on the growth performance and meat quality of Muscovy ducks.

MATERIAL AND METHODS

Experimental Design

A total of 800 male Muscovy ducks with an initial weight of (50.15 ± 5.37 g) was selected and randomly divided into 4 groups, with 4 replicates in each group, with 50 ducks in each replicate (each consisting of 4 pens as replicates, with 50 ducks per pen). The control group drank deep well water, while the experimental group added 1%, 3% and 5% DDL (1.59×108 CFU/mL) to the water. The test period is 70 d. The isolation and identification of DDL (ZG19) was based on the Berger Bacterial Identification Manual (9th Edition, Baltimore: Williams & Wilkins), and the fermentation and proliferation were carried out with reference to our previous fermentation conditions in the laboratory (Li et al., 2024). The basic diet is a full-price compound feed prepared based on the nutritional requirements of Muscovy ducks from 1 to 70 days old in Nutrient Requirements of Poultry (Dale, 1994). The specific basal diet composition and nutritional levels are shown in Table 1. The determination of crude protein and dry matter is in accordance with the national standards GB/T 6432-2018 and GB/T 6438-2007; the determination of calcium and total phosphorus is in accordance with the national standards GB/T 6436-2018 and GB/T 6437-2018. Diet metabolizable energy (MJ/kg) = [total energy consumed (MJ) - fecal energy (MJ)]/diet intake (kg) (Irawan et al., 2022). The trial period is 75 d, including a 5-d pretest period. The animal care and use protocol was approved by the Institutional Animal Care and Use Committee at Institute of Animal Husbandry and Veterinary Medicine of Fujian Academy Agricultural and Sciences(202307FJ010).Table 1 Basal dietary composition and nutritional level (air-dry basis).

Table 1Items	Starter diet(1–35 d)	Grower diet (35–75 d)	
Ingredients			
 Corn	44.6	42.5	
 Soybean meal (45% CP)	23.0	22	
 Wheat bran	19.6	22.16	
 Soybean oil	2.0	0	
 Limestone	1.2	1.1	
 CaHPO₄	1.2	1.4	
 NaCL	0.4	0.84	
 Rice bran meal	6	8	
 Premix1	2	2	
 Total	100.0	100	
Nutrient levels²			
 ME/(MJ/kg)	2.850	2.800	
 DM %	85.76	86.24	
 CP %	18.62	16.28	
 Ca %	1.17	1.19	
 AP %	0.33	0.32	
 Met %	0.49	0.47	
 Lys %	1.15	0.85	
 Try %	0.22	0.21	
 Thr %	0.78	0.56	
1 The premix provided the following per kg of diets: VA 12000IU;VD3 3600 IU;VE 20IU;VK3 2.4 mg ;VB1 2.0 mg;VB2 9.0 mg; D-pantothenic acid 12.0 mg;nicotinic acid 60.0 mg;VB6 4.2 mg;VB12 30 μg;biotin 0.15 mg; folic acid 1.5 mg; Fe (as ferrous sulfate) 36.0 mg; Cu (as copper sulfate) 4.0 mg; Zn (as zinc sulfate) 37.5 mg; Mn (as manganese sulfate) 45.0 mg; I (as potassium iodide) 0.25 mg; Se (as sodium selenite) 0.18 mg.

2 Nutrient levels was a calculated value, while the others were measured values.

Feeding Management

Muscovy ducks were raised in a mesh enclosure and provided with adlibitum feeding in the same facility. The temperature inside the duck house was maintained at approximately 35°C at the duckling, while the ambient temperature for the young ducks was kept at 25°C. Furthermore, standard procedures for immunization, deworming, health care, and disinfection was followed for the experimental ducks in line with the practices of the duck farm.

Sample Collection

Blood

On the final morning of the experiment, 4 Muscovy ducks from each experimental group were euthanized by inhaling carbon dioxide and cervical dislocation, performed by competent personnel who experienced and correctly applied the technique. and 5 mL of blood was extracted from the wing veins. The blood samples were collected and left at room temperature for 30 min before being centrifuged at 3000 g/min for 20 min. The supernatant was then stored at -20°C for future use in the next step. Additionally, other tissues and meat were gathered for subsequent experiments, the specifics of which are outlined below.

Intestine and Muscle Tissue

Four Muscovy ducks were selected to collect approximately 2 cm segments of the duodenum, jejunum, ileum, colon, and cecum. These segments were subsequently fixed in 10% paraformaldehyde, dehydrated, cleared, immersed in wax, embedded, and cut using a microtome for future analysis. The processed samples were then stored at -4°C for later examination.

INDICATOR MEASUREMENTS AND METHODS

The Assay of Growth Performance

Muscovy ducks were weighed after fasting on the 1st, 35th, and 75th d of the experiment, and their body weights were recorded. Throughout the experiment, the average feed intake (ADFI) was measured and calculated. At the end of the experiment, Muscovy ducks in both the experimental and control groups were weighed. Subsequently, the average daily gain (ADG) and feed-to-weight ratio (F/G) for all 4 groups of Muscovy ducks were computed using the following formulas: ADG (kg/d) = (final weight - initial weight) / number of experimental days; ADFI (kg/d) = total feed intake / number of experimental days; F/G = ADFI / ADG.

Assessment of Sensory, Amino Acids, and Fatty Acids of Duck Meat

Drawing from existing literature on sensory evaluation methodologies for meat and soup, a panel of 10 extensively trained reviewers (comprising 5 males and 5 females aged between 20 and 40) was carefully selected. The cooked duck meat and soup from both the experimental and control groups were divided into 10 equal portions, which were then placed in clean glass jars and sealed to maintain typical serving temperatures. The origin of the duck meat was kept undisclosed during the tasting sessions, ensuring that judges remained unaware of the group to which each sample belonged. Prior to the evaluation, judges were given specific instructions to abstain from communication, refrain from wearing makeup or carrying scented items, and rinse their mouths between tastings to avoid flavor contamination. The assessment criteria encompassed aspects such as meat flavor, sweetness, and elasticity, along with the umami and sweetness of the soup. Judges were required to rate the duck meat on a scale ranging from light to strong, assigning 1 point for absence, 3 points for mild, and 5 points for strong intensity. Professional sensory assessors then evaluated both the duck meat and soup using the same scoring system (1, 3, and 5 points), calculated the average scores, and determined the taste evaluation score for each sample by averaging the scores of the 3 indicators and dividing by 10. In the evaluation of aroma, judges were instructed to inhale gently, moderately, and forcefully, rinse their mouths between samples, and allow a 5-min interval before proceeding to the next assessment.

Refer to the National Food Safety Standard GB 5009.124-2016 for the determination of amino acids in foods. Hydrolyze duck meat using hydrochloric acid and analyze the amino acid content (Library., 2015). The fatty acids in duck meat can be determined to use the method outlined in AOAC 996.06 for the determination of fat content (Meinholz et al., 2008).

Determination of Blood Biochemical and Immune and Antioxidant Indicators of Muscovy Duck

Total protein (TP), albumin (TPT), glucose (GLU), urea nitrogen (UN), urea (U), globulin (GLO), and uric acid (UA) in serum was detected using an automatic blood biochemical analyzer (Beckman AU5800, Miami). UA, triglycerides (TG), total cholesterol (TC), high-density lipoprotein (HDL), and low-density lipoprotein (LDL). The blood immune indicators immunoglobulin G (IgG), albumin-globulin ratio (A/C), immunoglobulin M (IgM), and immunoglobulin A (IgA) were purchased from Tiangen Biotechnology (Beijing) Co., Ltd. Indicators of antioxidant capacity: malondialdehyde (MDA), total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) were purchased from Shanghai Sangon Bioengineering Technology Service Co., Ltd., and the determination method was performed according to the instruction manual of the kit.

Analysis of the Immune Organ Index

Four Muscovy ducks (the ducks from which blood was collected previously) of thymus, bursa and spleen were collected, and the excess water and blood were absorbed with filter paper, weighed and analyzed to calculate the organ index:Immuneorganindex(mg/g)=immuneorganweight(mg)/livingbodyweight(g)

Hematoxylin-Eosin, Tunel and Periodic Acid-Schiff (ab-pas) Staining and Analysis

The paraffin sections of each intestinal tissue prepared previously were dewaxed sequentially. After treatment with conventional hematoxylin-eosin staining (HE)), dehydration and gum mounting were performed. The gross structure of the bowel in each group was observed under a light microscope with a power of 100, and the morphology and distribution of the intestinal mucosa, submucosa, muscular layer and serosa in each group was observed under a light microscope with a power of 400. The height of villi and the depth of crypts were measured using DigiLab-C software. Five fields were selected for each section, and the average value was taken as the final result. The data were recorded and used to calculate the height of villi/crypt depth.

Tunel straining (duodenum and ileum were selected): the paraffin tissue sections should be placed in a staining vat and washed twice with xylene and absolute ethanol. Subsequently, they should be washed once with 95% ethanol and then with 75% ethanol. Following this, PBS containing proteinase K solution (20 µg/ml) should be added to hydrolyze the tissue proteins at room temperature. The sections should then be washed 4 times with distilled water. Next, they should be placed in a color tank with PBS containing 2% hydrogen peroxide for 2 washes. Subsequently, the TdT enzyme should be used to react. The sections should then be stained in a staining vat, incubated with peroxidase-labeled anti-digoxigenin antibodies, and stained with the methyl green. Finally, the sections should be sealed, dried, and observed under a light microscope to record the experimental results.

AB-PAS staining protocol was followed for tissue slices (duodenum and ileum). The slices underwent a series of sequential treatments: xylene I for 20 min, xylene II for 20 min, absolute ethanol I for 10 min, absolute ethanol II for 10 min, 95% alcohol for 5 min, 90% alcohol for 5 min, 80% alcohol for 5 min, and 70% alcohol for 5 min, followed by a rinse with distilled water. Subsequently, the slices were immersed in periodic acid solution for 10 min and rinsed with running water for 5 min. Next, the sections were treated with Schiff's reagent for 10 min, followed by a 5 min rinse with running water. The sections were then stained with hematoxylin for 1 min and rinsed with running water for 5 min. Finally, the slices underwent a dehydration process by being placed in 95% alcohol I for 5 min, 95% alcohol II for 5 min, absolute ethanol I for 5 min, absolute ethanol II for 5 min, xylene I for 5 min, and xylene II for 5 min to achieve transparency. The slices were then removed from xylene, slightly dried, and sealed with neutral gum. Subsequently, microscopic examination was conducted to capture images for further analysis. Statistical analysis was performed using SPSS 19.0 to analyze the significance of the difference among the groups and the changes in acidic mucus, neutral mucus, and glycogen.

16S Sequencing

Intestinal contents were aseptically collected from the duodenum and ileum of Muscovy ducks. The samples were then placed in a PBS buffer with a pH range of 7.2 to 7.4, along with a single steel bead, and gently shaken in an ice bath to disrupt the tissue and contents. Total DNA extraction was carried out using the TIANamp Stool DNA Kit, and the concentration and purity of the extracted DNA were evaluated with a spectrophotometer. Upon meeting quality standards, the DNA was stored at -20°C for further analysis. The V3 to V4 region of the 16S rDNA of intestinal bacteria was amplified through 16S rDNA sequencing using Illumina technology, with specific primer sequences being forward primer 341F (5′-CCTACGGNGGCWGCAG-3′) and reverse primer 805R (5′-GGACTACHVGGGTWTCTAAT-3′). After purification and quantification of PCR products, analysis was done using a bioanalyzer and Illumina's library quantification kit. Libraries with concentrations exceeding 2nM were appropriately diluted, mixed, denatured, and sequenced. Subsequent data processing involved obtaining RawData, followed by pairing, splicing, quality control, and chimera filtering to produce high-quality CleanData. DADA2, a method that no longer relies on clustering based on sequence similarity, was used, employing steps like ‘Dereplication’ to ensure single-base accuracy in representative sequences. This approach enhances data accuracy and species resolution. Amplicon sequence variants (ASV) were utilized to generate an operational taxonomic units (OTU) table, resulting in the final ASV feature table and feature sequences.This enables further analysis such as diversity assessment, species classification annotation, and differential analysis.

Statistical Analysis

The test data were analyzed by one-way analysis of variance (ANOVA) using SPS 19.0. The intergroup comparisons were performed using a t test. The test results are expressed as the mean ± standard error, with P > 0.05 indicating that the difference was not significant and P < 0.05 indicating that the difference was significant. And the correlation analysis was performed with Karl Pearson coefficient.

RESULTS

Effect of DDL Drinking on the Growth Performance and the Sensors of Meat in Muscovy Ducks

As shown in Table 2, compared with the control group, the DDL drinking group had no significant difference in initial weight, final weight or daily gain (P > 0.05) up to 35 d after the experiment. The ADFI decreased significantly (P < 0.05). ADFI of the fermented diet group supplemented with 5% DDL drinking was 7.87 g lower than that of the control group, and the difference was significant (P < 0.05). The feed-to-weight ratio also decreased significantly (P < 0.05). Compared with the control group, the feed to weight ratio and ADFI decreased significantly after the 75th d of the experiment, with the most significant decrease in the group supplemented with 5% DDL (P<0.05). The average daily gain and final weight between groups was not significantly different (P > 0.05).Table 2 Effects of DDL on the growth performance of Muscovy ducks.

Table 2Days	Items	Control group	1%	3%	5%	P-value	
35	Initial weight/g	50.43 ± 3.27	50.18 ± 4.39	50.15 ± 2.35	50.03 ± 3.58	0.563	
	Final weight/g	1134.16 ± 3.59	1091.34 ± 5.04	1088.08 ± 7.32	1118.01 ± 6.05	0.672	
	ADG/g	30.87 ± 3.36	29.75 ± 2.54	29.66 ± 3.29	30.43 ± 1.75	0.598	
	ADFI/g	65.45 ± 7.32a	60.31 ± 7.05b	58.56 ± 7.21b	57.58 ± 5.01b	0.034	
	F/G	2.11 ± 0.18a	2..03 ± 0.17b	1.97 ± 0.13b	1.88 ± 0.15b	0.047	
75	Initial weight/g	1134.16 ± 3.59	1091.34 ± 5.04	1088.08 ± 7.32	1118.01 ± 6.05	0.761	
	Final weight/g	2207.16 ± 3.59	2204.34 ± 5.04	2205.54 ± 7.32	2209.01 ± 6.05	0.633	
	ADG/g	30.65 ± 5.13	31.81 ± 4.17	31.93 ± 2.79	31.17 ± 2.53	0.525	
	ADFI/g	112.35 ± 11.32a	109.54 ± 15.90b	108.32 ± 10.43b	105.15 ± 15.07b	0.029	
	F/G	3.66 ± 0.13a	3.44 ± 0.112b	3.39 ± 0.15ab	3.37 ± 0.16ab	0.015	
Note: Values are mean ± SE, n = 10.

The different lower case letters in the table are significant (P < 0.05), and the same or no letters are not significant (P > 0.05). The following tables are the same.

Ten professional food sensory reviewers utilized the tasting method to evaluate the taste of cooked duck meat samples. The results, detailed in Table 3, indicate that DDL drinking group resulted in improved richness and sweetness in both the duck meat compared to the control group. Particularly, the duck meat from the 5% drinking group exhibited the highest levels of richness and sweetness (P < 0.05). Furthermore, the umami and tenderness evaluation of the duck meat revealed no significant differences among the 4 drinking water methods.Table 3 Effects of DDL on sensory of Muscovy duck meat.

Table 3	The proportion of DDL in water %		
Items	(Control)	1	3	5	P-value	
Richness	3.23 ± 0.15c	3.59 ± 0.19b	4.25 ± 0.24a	4.78 ± 0.42a	0.019	
Sweetness	3.02 ± 0.27c	4.01 ± 0.14b	4.17 ± 0.09b	4.30 ± 0.05a	0.034	
Tenderness	3.56 ± 0.33	3.25 ± 0.18	3.21 ± 0.51	3.42 ± 0.25	0.377	
Umami	3.35 ± 0.23	3.27 ± 0.21	3.45 ± 0.09	3.69 ± 0.08	0.562	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05).

Effect of DDL Drinking on Muscovy Duck Serum Biochemical Indicators

As shown in Table 4, DDL drinking could significantly increase the A/G ratio (P < 0.05). The TG and TC contents in serum were increased (P < 0.05). The contents of TG and TC in the serum of the groups supplemented with 3% and 5% DDL drinking were significantly higher than those in the control group and 1% group (P < 0.05). Other serum biochemical indicators were not significantly different (P > 0.05).Table 4 Effects of DDL on biochemical indexes of Muscovy duck serum.

Table 4	The proportion of Lactobacillus from duck %		
Items	(Control)	1	3	5	P-value	
GLU/(mmol/L)	6.91 ± 0.87	7.11 ± 0.83	7.01 ± 0.19	7.21 ± 0.61	0.439	
TP/(g/L)	43.15 ± 5.54	42.55 ± 8.90	45.12 ± 7.53	46.30 ± 7.16	0.521	
ALB/(g/L)	13.11 ± 3.34	14.01 ± 2.76	15.05 ± 5.77	16.10 ± 4.34	0.128	
GLO/(g/L)	29.23 ± 3.98	30.04 ± 7.55	31.23 ± 8.30	29.65 ± 5.81	0.233	
A/G	0.38 ± 0.11c	0.47 ± 0.15b	0.51 ± 0.07a	0.55 ± 0.03a	0.011	
ALP/(U/L)	201.54 ± 25.31	199.32 ± 32.70	205.21 ± 32.56	203.50 ± 32.1	0.355	
UN/(mmol/L)	0.55 ± 0.01	0.61 ± 0.04	0.59 ± 0.03	0.63 ± 0.01	0.259	
UA/(mmol/L	132.12 ± 10.33	131.54 ± 9.76	132.61 ± 13.53	130.50 ± 11.35	0.220	
TG/(mmol/L)	0.36 ± 0.03c	0.59 ± 0.02b	0.75 ± 0.01a	0.81 ± 0.05a	0.037	
TC/(mmol/L)	3.13 ± 0.45c	3.79 ± 0.15b	4.67 ± 0.53a	4.75 ± 0.15a	0.011	
HDL/(mmol/L)	1.35 ± 0.13	1.38 ± 0.21	1.32 ± 0.50	1.37 ± 0.22	0.651	
LDL/(mmol/L)	1.37 ± 0.21	1.35 ± 0.21	1.31 ± 0.43	1.35 ± 0.56	0.349	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

Effects of DDL Drinking on Immune Indicators and Antioxidant Capacity of Muscovy Ducks

As shown in Table 5, DDL drinking could significantly increase the indices of the immune organs thymus and fabricius bursa of Muscovy duck (P < 0.05), among which 5% was the highest index. Compared with the control group, DDL drinking had an exponential trend of increasing the liver and spleen, but the difference was not significant (P > 0.05). Additionally, compared with the control group, the contents of IgG in the serum of the DDL group were significantly higher (P < 0.05), and 5% group was the most significant (P < 0.05) (Table 6). There was no significant difference in the concentration of IgA and IgM in the serum of the 4 groups (P > 0.05). Compared with the control group, the overall antioxidant capacity of DDL drinking was higher in terms of T-AOC, MDA and superoxide dismutase (SOD) (P > 0.05). The content of MDA in the serum of the Muscovy duck drinking with 5% DDL was the highest, and the difference was significant (P < 0.05). The levels of catalase CAT and glutathione peroxide GSH-Px in Muscovy ducks serum were not significantly different among the 4 groups (P > 0.05).Table 5 Effect of DDL on immune organ index of Muscovy ducks.

Table 5Items	Control group	1%	3%	5%	P-value	
Liver index	0.56 ± 0.07	0.58 ± 0.09	0.65 ± 0.11	0.63 ± 0.08	0.332	
Thymus index	3.16 ± 0.59c	4.34 ± 0.04b	4.54 ± 0.32b	5.01 ± 0.35a	0.017	
Fabricius index	1.34 ± 0.11c	1.33 ± 0.14c	1.16 ± 0.27b	1.67 ± 0.39a	0.023	
Spleen index	3.99 ± 0.24	4.07 ± 0.03	4.22 ± 0.02	4.27 ± 0.05	0.154	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

Table 6 Effect of DDL on immune and antioxidant indexes in Muscovy ducks.

Table 6Items	Control group	1%	3%	5%	P-value	
IgG/(mg/mL)	43.26 ± 4.87d	49.58 ± 6.53c	53.09 ± 6.74b	55.21 ± 3.95a	0.022	
IgM/(mg/mL)	4.25 ± 0.41	4.33 ± 0.29	4.59 ± 0.37	4.53 ± 0.10	0.251	
IgA/(mg/mL)	0.38 ± 0.12	0.39 ± 0.17	0.41 ± 0.08	0.40 ± 0.19	0.642	
T-AOC/(U/mL)	4.65 ± 0.95c	5.32 ± 0.77b	6.05 ± 0.59a	6.12 ± 0.30a	0.013	
MDA(nmol/mL)	3.27 ± 0.78c	3.36 ± 0.52c	3.79 ± 0.54b	4.31 ± 0.75a	0.015	
CAT(U/mL)	1.76 ± 0.12	1.83 ± 0.42	1.77 ± 0.29	1.84 ± 0.35	0.577	
T-SOD/(U/mL)	54.75 ± 3.97b	56.32 ± 6.73a	59.43 ± 7.65a	58.11 ± 6.55a	0.061	
GSH-Px/(U/mL	444.56 ± 13.25	459.30 ± 21.07	455.36 ± 13.67	457.26 ± 31.34	0.715	
Values are mean ± SE, n = 4.

a-d Values within a row with different letters differ significantly (P < 0.05)

Effects of DDL Drinking on Intestinal Morphology, Intestinal Mucosa and Intestinal Epithelial Cells of Muscovy Ducks

As shown in Table 7, compared with the control group, the height of the jejunum and colon villi in DDL drinking group was higher (P < 0.05). The crypt height of the ileum of the Muscovy duck in the DDL drinking group was decreased compared with that in the control group, and there was a significant difference (P < 0.05). The ratio of the villi height and the crypt depth in the jejunum and colon were significantly increased in the DDL drinking group (P < 0.05), and the V/C, villi height and the crypt depth ratio of the duodenum and ileum showed no significant difference between the groups (P > 0.05).Table 7 Effects of Lactobacillus fermentation on intestinal morphological structure of Muscovy ducks.

Table 7Items	Control	1%	3%	5%	P-value	
Duodenum						
 Villus height/μm	430.52 ± 5.70	435.51 ± 7.35	426.77 ± 9.61	435.32 ± 9.98	0.176	
 Crypt depth/μm	297.21 ± 5.76	295.36 ± 7.89	283.37 ± 8.05	291.33 ± 7.06	0.541	
 V/C	1.45 ± 0.12	1.47 ± 0.25	1.51 ± 0.24	1.49 ± 0.15	0.216	
Jejunum						
 Villus height/μm	345.61 ± 10.76c	359.30 ± 8.23b	379.51 ± 9.11a	378.44 ± 6.73a	0.017	
 Crypt depth/μm	180.17 ± 5.33	182.45 ± 6.18	183.56 ± 7.08	183.45 ± 6.55	0.432	
 V/C	1.91+0.14b	1.97+0.21b	2.07+0.77a	2.06+0.32a	0.026	
Ileum						
 Villus height/μm	255.31 ± 5.60	257.35 ± 6.31	249.37 ± 9.55	252.13 ± 5.02	0.076	
 Crypt depth/μm	131.21 ± 3.59a	133.54 ± 7.06a	122.10 ± 9.17b	125.22 ± 5.05b	0.042	
 V/C	1.95 ± 0.06	1.93 ± 0.12	2.04 ± 0.41	2.01 ± 0.39	0.598	
Colon						
 Villus height/μm	328.57 ± 9.99c	339.01 ± 8.57b	349.53 ± 10.93a	343.39 ± 7.54a	0.021	
 Crypt depth/μm	161.15 ± 6.43	166.53 ± 8.02	164.56 ± 8.23	164.57 ± 9.71	0.113	
 V/C	2.03 ± 0.06b	2.04 ± 0.15b	2.12 ± 0.40a	2.08 ± 0.56a	0.016	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

The PAS reagent binds to glycoproteins (neutral and acidic mucins) and lipids in the secreted mucus from the intestines, staining goblet cells in magenta. In Muscovy duck intestinal mucosa (Figure 1), DDL drinking significantly increased the ratio of glycoproteins and goblet cells on the mucosal surface of the ileum and duodenum compared to the control group. There was a significant difference in the number of goblet cells between the DDL drinking group and the control group (P < 0.05). While there was an increasing trend in the number of goblet cells on the mucosal surface of the ileum and duodenum in the DDL drinking group compared to the control group, this trend was not significant in the 3% DDL group (P > 0.05). The number of goblet cells on the duodenal mucosal surface did not show a significant difference between groups (P > 0.05).Figure 1 Effects of DDL drinking on mucus and glycoproteins in duodenum and ileum of Muscovy ducks. A Control. B 1% DDL drinking group. C 3% DDL drinking group. D 5% DDL drinking group. E and F the ratio of mucus, glycoproteins and tissue in duodenum and ileum. Data presented as means ± SD (n = 5). Significant are marked with asterisks: *0.01 <P ≤ 0.05; **0.001 < P ≤ 0.01.

Figure 1

TUNEL was a widely used approach for detecting DNA fragmentation during apoptosis. As shown from Figure 2, it was observed that DDL drinking led to a significant reduction in the number of Tunel-positive cells in the duodenum and ileum of Muscovy ducks, with the most pronounced decrease seen in the 5% DDL drinking group.Figure 2 Effects of DDL drinking on intestinal(duodenum and ileum) epithelial cells of Muscovy ducks. A Duodenum. B ileum. C Tunel positive cells. Data presented as means ± SD (n = 5). Significant are marked with asterisks: *0.01 <P ≤ 0.05; **0.001 < P ≤ 0.01.

Figure 2

Effect of DDL Drinking on the Meat Quality Composition of Duck

To further explore the impact of DDL drinking on enhancing duck meat quality, we analyzed the saturated and unsaturated fatty acids and amino acid components present in the meat. The findings in Table 8 revealed a significant increase in saturated fatty acids (C12:0) and unsaturated fatty acids (C18:1 n-9, C18:1 n-7, C18:2 n-6, C20:5 n-3(EPA), C22:5 n-3 (DPA), C22:6 n-3 (DHA), with the most pronounced effect observed in the 5% DDL drinking group. Furthermore, DDL drinking led to a notable rise in the levels of Glycine (Gly), Serine (Ser), Glutamic acid (Glu), and Tyrosine (Tyr) in the meat (breast), and Glutamic acid showing the most prominent increase, but other AAs was not significantly difference (Table 9).Table 8 Effects of DDL on fatty acid composition of breast muscle of the Muscovy ducks (mg/kg) (n = 4, slaughtered at 75 d of age).

Table 8	DDL		
Fatty acids	Control	1%	3%	5%	P-value	
C12:0	0.08 ± 0.02d	0.21 ± 0.03c	0.47 ± 0.02b	0.75 ± 0.01a	0.023	
C14:0	0.33 ± 0.01	0.35 ± 0.01	0.41 ± 0.01	0.45 ± 0.01	0.125	
C16:0	19.35 ± 1.02	19.78 ± 1.78	20.15 ± 2.13	19.67 ± 2.12	0.331	
C18:0	10.15 ± 0.87	11.54 ± 1.95	10.98 ± 2.13	10.58 ± 1.54	0.276	
C16:1 n-7	1.1 ± 0.12	1.15 ± 0.43	1.11 ± 0.18	1.09 ± 0.58	0.655	
C18:1 n-9	20.22 ± 3.45c	22.34 ± 2.72b	24.45 ± 3.56a	26.12 ± 1.89a	0.029	
C18:1 n-7	3.28 ± 0.25d	3.65 ± 0.34c	3.79 ± 0.17b	3.97 ± 0.51a	0.014	
C18:2 n-6	21.12 ± 3.15c	23.57 ± 4.16b	24.81 ± 1.65a	24.93 ± 2.77a	0.018	
C18:3 n-3	2.03 ± 0.07	2.04 ± 0.12	2.06 ± 0.08	2.09 ± 0.09	0.261	
C20:2 n-6	1.22 ± 0.04	1.27 ± 0.05	1.24 ± 0.01	1.29 ± 0.01	0.530	
C22:5 n-6	0.38 ± 0.01	0.39 ± 0.01	0.41 ± 0.02	0.41 ± 0.03	0.612	
C20:5 n-3 (EPA)	1.21 ± 0.05c	1.45 ± 0.01b	1.55 ± 0.01b	1.76 ± 0.02a	0.015	
C22:5 n-3 (DPA)	1.04 ± 0.01c	1.25 ± 0.02b	1.29 ± 0.03b	1.37 ± 0.02a	0.019	
C22:6 n-3 (DHA)	2.00 ± 0.11c	2.14 ± 0.033c	2.35 ± 0.01b	3.81 ± 0.41a	0.007	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

Table 9 Effects of DDL on amino acid composition of breast muscle of the Muscovy ducks (%).

Table 9	Control	1%	3%	5%	P-value	
	Breast	Breast	Breast	Breast		
Asparagic acid (Asp)	0.79 ± 0.01	0.88 ± 0.10	0.79 ± 0.02	0.79 ± 0.04	0.339	
Glycine (Gly)	0.92 ± 0.07b	0.97 ± 0.01b	0.99 ± 0.03b	1.21 ± 0.03a	0.008	
Alanine (Ala)	0.97 ± 3.21	0.97 ± 1.90	0.89 ± 0.03	0.98 ± 005	0.145	
Proline (Pro)	1.23 ± 0.14	1.21 ± 0.30	1.34 ± 0.15	1.22 ± 0.01	0.320	
Arginine (Arg)	0.89 ± 0.13	0.87 ± 0.12	0.82 ± 0.03	0.83 ± 0.05	0.125	
Serine (Ser)	1.35 ± 0.19c	1.33 ± 0.11c	1.52 ± 0.06b	2.01 ± 0.02a	0.029	
Cysteine (Cys)	0.75 ± 0.05	0.83 ± 0.11	0.79 ± 0.02	0.73 ± 0.01	0.112	
Glutamic acid (Glu)	1.11 ± 0.21c	1.56 ± 0.03c	2.92 ± 0.32a	2.75 ± 0.32a	0.035	
Threonine (Thr)	1.08 ± 0.03	1.03 ± 0.10	1.02 ± 0.04	1.02 ± 0.12	0.421	
Isoleucine (Ile)	0.77 ± 0.02b	1.01 ± 0.05a	1.21 ± 0.07a	1.08 ± 0.05a	0.022	
Leucine (Leu)	0.91 ± 0.09	1.05 ± 0.04	1.13 ± 0.12	1.08 ± 0.15	0.453	
Tyrosine (Tyr)	0.51 ± 0.01c	1.16 ± 0.25c	1.46 ± 0.32a	1.36 ± 0.33b	0.007	
Histidine (His)	0.65 ± 0.03	0.73 ± 0.22	0.62 ± 0.32	0.75 ± 0.01	0.105	
Phenylalanine (Phe)	1.02 ± 0.04	1.27 ± 0.03	1.40 ± 0.05	1.15 ± 0.14	0.209	
Methionine (Met)	1.28 ± 0.05	1.13 ± 0.23	1.13 ± 0.12	1.35 ± 0.34	0.322	
Valine (Val)	0.75 ± 0.01	0.85 ± 0.23	0.81 ± 0.30	0.73 ± 0.12	0.558	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

Effects of DDL Drinking on The Diversity of Intestinal Bacterial Communities in Muscovy Ducks

Alpha diversity analysis revealed a significant impact of lactic acid bacteria in DDL drinking group on the abundance of intestinal bacterial communities in Muscovy ducks (Table 10). The groups 3% and 5% DDL had the highest abundance of intestinal bacteria, as indicated by observed species (1033.13 ± 131.55) and chao1 (1127.11 ± 164.25) metrics. In contrast, the control group exhibited lower bacterial abundance (observed species: 910.35 ± 140.69, chao1: 857.41 ± 108.22). Furthermore, the diversity index of intestinal bacterial communities in the lactic acid bacteria drinking groups was higher than that of the control group, with the 5% DDL group showing the highest diversity index (Shannon: 5.96 ± 0.31, Simpson: 0.87 ± 0.03). Subsequently, a taxonomic analysis of the impact of DDL drinking group on bacterial communities in duck intestines was conducted. The results demonstrated significant differences between the DDL drinking group and the control group. Specifically, DDL drinking group exhibited higher abundance of Bacillus, Lentilactobacillus, and Bacteroides in the duodenum, while the control group had higher levels of Mictochondria. In the ileum, the most abundant bacterial genera was Lactobacillus, Ligilactobacillus, and Bacteroides, with higher levels of Enterobacter observed in the control group (Figure 3). These differences were statistically significant.Table 10 Effects of DDL on intestinal microbiota of Muscovy ducks.

Table 10Items	Control group	1%	3%	5%	P-value	
Species	910.35 ± 140.69c	1005.11 ± 103.27b	1033.1 ± 131.55a	1055.29 ± 191.26a	0.025	
Chao1	857.41 ± 108.22c	1006.14 ± 112.03b	1025.59 ± 103.64b	1127.11 ± 164.25a	0.031	
Shannon	3.83 ± 1.12c	4.75 ± 0.42b	5.77 ± 0.51a	5.96 ± 0.31a	0.032	
Simpson	0.70 ± 0.01c	0.77 ± 0.02b	0.75 ± 0.01b	0.87 ± 0.03a	0.018	
Values are mean ± SE, n = 4.

a-c Values within a row with different letters differ significantly (P < 0.05)

Figure 3 Changed the gut microbiota at the genus level after DDL drinking. A relative abundance of the gut microbiota at the genus level of 5% group DDL drinking group(Duodenum). B-D Relative abundance of bacillus, lentilactobacillus, bacteroides and mitochondria at the Duodenum(5% DDL drinking group). F Relative abundance of the gut microbiota at the genus level of 5% group DDL drinking group (ileum) G-I Relative abundance of lactobacillus, enterobacter, ligilactobacillus and bacteroides at the Duodenum. Data represent the Mean±SEM (n = 4). Mean values with different letters over the bars are significantly different (P < 0.05) according to a Duncan's test.

Figure 3

Correlation Between the Various Influencing Factors of Improved Duck Growth Performance and the Meat Composition

The Pearson correlation coefficient is a metric used to quantify the relationship between 2 variables, ranging from -1 to 1. A value of 1 indicates a perfect positive correlation, 0 signifies no linear correlation, and -1 represents a perfect negative correlation. To investigate the impact of DDL drinking group on duck production and meat quality, we utilized the Karl Pearson coefficient. Figure 4 findings revealed that the increased abundance of Ligilactobacillus and Lactobacillus in the ileum, due to DDL consumption, was associated with elevated levels of unsaturated fatty acids (C18:1n-7, C20:2 n-6, C22:5), and serine in the meat. These factors exhibited positive correlations, while negative correlations were observed with duck's F/G ratio. Furthermore, the increase in lentibacillus beneficial bacteria in the duodenum, induced by DDL intake, showed positive correlations with unsaturated fatty acids (C20:5 n-3, DPA) and glutamic acid in the meat, enhancing its sweetness and flavor. Additionally, the growth of villi in the duodenum and ileum post lactic acid consumption was negatively correlated with F/G ratio. Notably, a positive correlation was observed between IgA levels in blood immunoglobulins and the immune index after lactic acid bacteria ingestion.Figure 4 Pearson's correlation cluster heatmap of meat sensor, meat ingredients, immune indicators and antioxidant capacity, Gut microbiota, growth performance, and gut morphology of Muscovy duck after DDL drinking intervention. Darker red and darker blue represents higher levels of positive and negative correlations, respectively. Significant correlations are marked with asterisks: *0.01 < P ≤ 0.05; **0.001 < P ≤ 0.01.

Figure 4

DISCUSSION

Effect of DDL Drinking on the Growth Performance of Muscovy Duck

DDL is a beneficial bacteria, which can significantly improve the growth performance of poultry, increase the daily gain, reduce the feed-to-weight ratio, and increase the economic benefits of poultry farming (Reuben et al., 2019, Li et al., 2024). Previous studies reported that feeding fermented feed with probiotics to broiler chickens could significantly increase the feed intake and daily gain and reduce the feed-to-weight ratio (Zhang et al., 2024). Another study reported that Cherry Valley ducks fed DDL fermented feed could also significantly increase their daily gain and feed-to-weight ratio, but there was no significant difference in average daily feed intake (Zhang et al., 2020). Previous our research revealed that Muscovy ducks feed with the probiotics (DDL and bacillus) fermented diet significantly improved the growth performance by shaping the gut morphology, microbiota and metabolism (Li et al., 2024). The results of the present study showed that ADFI and feed-to-weight ratio of Muscovy duck was significantly decreased when DDL drinking was lasting for 35 d, while the average daily gain of Muscovy duck had no significant change. While Muscovy ducks drink DDL to 75 days old, ADFI and feed-to-weight ratio continued to decline. In addition, in this study, as the concentration of drinking DDL increases, the daily feed intake and feed-to-weight ratio of the experimental group decreased significantly, while the daily weight gain showed no significant difference. It may be that after DDL drinking promotes the decomposition and metabolism of the feed and improves the gastrointestinal digestion and the absorption rate of crude protein, crude fiber and neutral detergent fiber in the feed. However, the specific and detailed mechanism remains to be studied in depth in the future.

Effect of DDL Drinking on Muscovy Duck Serum Biochemical Indicators

The detection of biochemical indicators in Muscovy duck serum could reflect the protein metabolism and fat metabolism of Muscovy duck in real time (Qi et al., 2024). The concentration of total proteins in serum can effectively reflect the strength of protein anabolism in the body (Zhuang et al., 2022). This study found that DDL drinking significantly increased the total protein concentration of duck serum, promoted the deposition of more proteins in the tissues of Muscovy duck, and accelerated the growth and development of the tissues and organs of Muscovy duck. The result was that the growth performance of Muscovy duck was improved. Cholesterol and triglycerides in serum is indicators of the body's lipid metabolism function, and sufficient amounts of cholesterol and triglycerides are key for the growth and development of the body (Zhu et al., 2020). This study found that DDL drinking group significantly increased the cholesterol and triglyceride levels in serum, with the 3% and 5% DDL drinking groups having the most significant increase. These results suggest that DDL drinking can promote fat metabolism and synthesis and improve the growth performance of Muscovy duck.

Effects of DDL Drinking on the Immune Indicators and Antioxidant Capacity of Muscovy Ducks

Blood immune indicators can reflect the immune status of Muscovy ducks (Liu et al., 2021). IgG plays an important role in the humoral immune response and plays a key role in the control of bacterial infections in the body. IgA is the mucosal immunity antibody second only to IgG and a key of mucosal immunity of the body and is mainly produced by the mucosal epithelium of the gastrointestinal tract. It can bind to a variety of pathogenic antigens at critical moments to control the diarrhea infection. Previous studies reported that the concentrations of IgG and immunoglobulin M (IgM) in the blood of chickens, geese and Cherry Valley ducks were significantly increased after a period of feeding probiotics (Zhu et al., 2015). It has been reported that the serum IgG level of meat ducks fed probiotics fermented feed was lower, and there was no difference in the serum levels of immunoglobulins IgA and IgM (Qiu et al., 2022). The results of this study showed that the levels of IgG and IgM in the blood of DDL were significantly increased, with the increase in IgG being the most significant, while IgA had no significant change. Previous studies have shown that fatty acids, including monounsaturated fatty acids (MUFAs), polyunsaturated fatty acids (PUFAs) and SFAs, not only improve meat quality but also have regulatory effects on the body's hormone metabolism and the activities of various enzymes. It plays a wide range of roles in lipid metabolism, prevention of cardiovascular and cerebrovascular diseases, and delaying immune function decline (Fu et al., 2023). Therefore, the significant increase in IgG and IgM in this study may be closely related to the large amount of fatty acids produced by microbial fermentation.

Free radicals are often criticized as a synonym for aging and disease in the biological field. Therefore, as an indicator of the body's ability to scavenge free radicals, the total antioxidant capacity is often used as one of the indicators of body health (Szabó et al., 2023). Among them, GSH-Px, T-SOD and CAT are the most important enzymatic antioxidant systems in the body, and their main functions are to remove excessive free radicals and reactive oxygen species from the body and prevent the production of lipid peroxides (Duangnumsawang et al., 2021). This study found that DDL drinking significantly increased the T-OA contents of Muscovy ducks, and then improved the level of antioxidant capacity.

Effect of DDL Drinking on the Intestinal Morphology of Muscovy Ducks

Research has shown that probiotics fermented feed can alter the morphology of the gastrointestinal tract, leading to a significant increase in villi length and a higher villi/crypt ratio when fed to poultry (Liu et al., 2023a). The small intestine villi in chickens and ducks are the fastest-growing tissue and play a crucial role in absorbing essential nutrients for growth directly from the intestines. In instances of short-term 'starvation' of the small intestine, there is a rapid decrease in villi length, which hampers small intestine growth. The absorption of nutrients in the intestines has a direct impact on growth and development (Zhu et al., 2020). The DDL drinking method utilized in this study addressed the “starvation” state of the small intestine caused by poor solid feed palatability, providing necessary nutrients for Muscovy ducks, sustaining the growth of intestinal villi, enhancing nutrient absorption efficiency, and ultimately promoting overall body growth and development.

Effect of DDL Drinking on the Meat Quality of Muscovy Ducks

Meat tenderness, nutritional composition, amino acid, and fatty acid content are key factors influencing meat taste (Liu et al., 2018). Tenderness impacts taste, while nutrients and acids determine flavor. Amino acids and fatty acids are fundamental flavor components in meat, directly influencing its taste. Lactic acid bacteria, as beneficial microbes, have a significant impact on the quality of meat products from livestock and poultry. Research indicates that supplementing diets with LAB can mitigate hydrogen peroxide production by intestinal microorganisms in poultry, reduce cholesterol levels, enhance the meat color, decrease the meat loss, and increase muscle fat content (de Souza et al., 2022). Feeding animals probiotics containing lactobacilli can boost serum lipase activity, leading to fat breakdown into fatty acids and glycerol (Eglite et al., 2023). Studies on broiler chickens fed lactic acid bacteria have shown a substantial increase in crude fat content in breast muscles compared to control groups (Grootjans et al., 2013). The ratio of oleic acid/linoleic acid in muscle fat directly impacts the meat flavor. LAB supplementation can also elevate fatty acid levels in muscles, with significantly higher levels of palmitoleic acid compared to the control groups (P < 0.05). Notably, the treatment with DDL drinking resulted in significantly higher levels of arachidonic acid (C20:2), arachidonic acid (C20:3), and polyunsaturated fatty acids were crucial for duck meat flavor formation. Studies have shown that supplementing meat poultry diets with 3 g/kg of compound probiotics containing LAB, such as Lactobacillus, Bacillus, Streptococcus faecalis, and yeast, can significantly increase the ratio of unsaturated fatty acids to saturated fatty acids in leg and breast muscles, as well as the linolenic acid content (Liu et al., 2023b). Continuous consumption of LAB by meat ducks was found to notably enhance the levels of unsaturated and monounsaturated fatty acids in the meat. This finding aligns with previous research indicating that adding 2% and 3% of complex probiotics with LAB to broiler diets can improve broiler chicken meat quality (Wu et al., 2022). Furthermore, essential amino acids like threonine, glutamic acid, alanine, lysine, and cysteine are crucial for the meat flavor. Studies have demonstrated that supplementing feed with LAB can significantly boost the accumulation of amino acids in meat, particularly those essential for flavor (Zhu and Yang, 2020). The present study revealed that the DDL drinking group significant increase in glutamic acid content in the meat, which may explain why previous studies reported improved sweetness and taste of duck meat after DDL consumption compared to a control group. Additionally, the correlation analysis confirmed a positive relationship between consuming LAB, meat sweetness and taste, and glutamic acid content in duck meat.

Effect of DDL Drinking on the Intestinal Glycoproteins (Neutral and Acidic Mucins) and Lipids and Goblet Cells of Muscovy Ducks

Intestinal glycoproteins and goblet cells in the intestinal mucosa are typical mucus cells that primarily consist of mucin. These mucin granules are released from the cells and produce mucus containing water and inorganic salts, which adhere to the intestinal mucosal surface, forming a protective layer of intestinal mucus that provides lubrication (Liu et al., 2020). The findings of this study demonstrated that at 75 d of age, the number of glycoproteins and goblet cells in the duodenum and ileum of the DDL drinking group was significantly higher compared to the control group; the increase in intestinal mucosal goblet cells in the 3% supplemented group was more pronounced. This could be attributed to the fact that DDL drinking can secrete a variety of beneficial small molecules, such as amino acids, vitamins, active peptides, digestive enzymes, and organic acids, which help protect the intestinal mucosa, enhance the growth of small intestinal villi, and facilitate the proliferation of beneficial microorganisms. These microorganisms in the grains can work together with mucin as an intestinal mucosal barrier to prevent pathogenic bacteria from entering the body, while also regulating the production of goblet cells and mucin content in intestinal secretions to maintain a stable number of glycoproteins and goblet cells in the intestine, thereby preserving the structural integrity of the intestinal tract (Rose et al., 2021). Furthermore, some small molecules and beneficial metabolites present in fermentation products can easily traverse the mucus layer, enhancing their absorption by the body.

CONCLUSIONS

In this study, drinking with DDL enhanced the body's immunity and antioxidant function, improved the structure of intestinal villi, increased the number of glycoproteins and goblet cells in the mucosa, up-regulated the absorption area of the intestinal mucosa, and promoted the response of the small intestine to nutrients substance absorption, and then improved the growth performance of Muscovy ducks. Moreover, DDL drinking significantly elevated the meat quality by increased the unsaturated fatty acids and amino acids related to taste.

ETHICS APPROVAL AND CONSENT TO PARTICIPATE

All the procedures were approved by Institutional Animal Care and Use Committee at Institute of Animal Husbandry and Veterinary Medicine of Fujian Academy Agricultural and Sciences (202307FJ010).

DISCLOSURES

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Appendix Supplementary materials

Image, application 1

ACKNOWLEDGMENTS

We thank all the members of the Laboratory for their support and constructive comments. This work was partially supported by the Fujian Provincial Spark Project (2022S0011 ), Fujian Provincial Competitive Public Welfare Project (2023R1076 ), Key Scientific and Technological Project of Fujian Academy of Agricultural Sciences (KJZD202404 ), 2022 "Tianshan Talents" Training Program of Xinjiang Uygur Autonomous Region-Young Top Science and Technology Talents Project (2022TSYCJC0022).

Supplementary material associated with this article can be found in the online version at doi:10.1016/j.psj.2024.104195.
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