Use of Autochthonous Lactic Acid Bacteria as Probiotic Additives for Muscovy Ducks in Housing

ABSTRACT

This study aimed to investigate the effects of autochthonous lactic acid bacteria (LAB) as probiotic additives for Muscovy ducks on performance, carcass traits, and serum biochemical parameters. LAB were isolated from the intestinal content of 12 Muscovy ducks, and three promising strains were identified: Enterococcus lactis, Enterococcus ratti, and Enterococcus faecium. Ninety-six male Muscovy ducks with eight days of age (weight = 158.56±2.17) were distributed in a completely randomized design, where the treatments comprised a control group and three experimental groups subjected to autochthonous LAB as a probiotic (E. lactis, E. ratti, and E. faecium) administered orally to the Muscovy ducks, with three replicates of eight birds each. Birds were monitored over 90 days, divided into starter, grower, and finisher stages. Blood was collected at 91 days of age for analysis, and at 91 days birds were slaughtered to evaluate carcass traits. Results indicated no significant effect (p>0.05) in feed intake and weight gain during the starter stage, though feed conversion ratio worsened (p<0.05) with probiotics. In the grower stage, E. lactis improved (p<0.05) feed conversion ratio. In the finisher stage and overall performance, the control group presented better (p<0.05) results. Carcass analysis showed E. faecium increased carcass and breast yield, but all probiotic groups had lower (p<0.05) slaughter weight results. Serum biochemical analysis revealed E. faecium influenced triglycerides, glucose, cholesterol, and albumin levels, suggesting metabolic changes. In conclusion, autochthonous LAB had varied effects on Muscovy duck performance and physiology, indicating that probiotic efficacy depends on the developmental stage and specific strains used.

Keywords:
Additive; Cairina moschata; feed conversion ratio; lactic acid bacteria; probiotic

INTRODUCTION

Muscovy ducks represent an important segment in the poultry industry, supplying various products to the market such as meat, eggs, viscera, feathers, and fatty livers known as foie gras (Islam et al., 2020; Santos et al., 2020; Arias-Sosa & Rojas, 2021). They cater to several important consumer markets, mainly located in Europe and Asia. However, they remain relatively unexplored in Latin America, despite their origin being directly related to this region of the world (Ardiansah et al., 2020; Jalaludeen et al., 2022). In this sense, it is also important to mention that, despite their market potential, Muscovy duck production remains relatively small compared to other poultry farming segments (Rufino et al., 2017a,b; Islam et al., 2020; Arias-Sosa and Rojas, 2021), representing a modest share within the Brazilian poultry production chain (Pingel, 2011; Santos et al., 2020; ABPA, 2023).

Among the nutritional alternatives to improve the production of Muscovy ducks, especially in housing, probiotics stand out for their potential to improve intestinal health at structural and microbial levels, which can induce better digestion, absorption, and nutrient utilization (Haghighi et al., 2005; Jeni et al., 2021). Additionally, they can enhance the health of intestinal microbiota, consequently inhibiting the action of pathogenic enteric microorganisms and improving performance (Rescigno et al., 2001; Haghighi et al., 2005; Jeni et al., 2021).

It is important to mention that the use of probiotics as additives in the poultry industry has been increasing, with autochthonous bacteria (also known as native microorganisms) garnering significant attention due to their ability to naturally colonize the host’s gastrointestinal tract among birds (Butel, 2014; Qiu et al., 2022). These bacteria are inherently adapted to the specific conditions of the host’s environment, which can enhance their efficacy in promoting gut health, improving nutrient absorption, and boosting overall performance (Qiu et al., 2022). This approach aligns with sustainable and natural farming practices, reducing the reliance on antibiotics and contributing to the production of healthier poultry products (Jeni et al., 2021).

The literature reports some strains of microorganisms that show potential to be used as probiotics, with the most common ones being lactic acid bacteria (LAB) and bifidobacteria (Didari et al., 2014; Jeni et al., 2021), especially strains from the genera Lactobacillus spp., Bifidobacterium spp., Enterococcus spp., Bacillus subtilis, and Sacharomyces spp. (Gaggia et al., 2010; Alagawany et al., 2018). It is also important to highlight that LAB gain significant attention as probiotics in the poultry industry because they are strains that thrive in gastrointestinal adversities, providing beneficial effects as performance enhancers and promoters of antagonism against enteric pathogens (El-Sawah et al., 2020).

In this context, it was hypothesized that autochthonous LAB, that is, bacteria isolated directly from the gastrointestinal tract of Muscovy ducks, could be used as probiotic additives for Muscovy ducks in housing due to their potential to enhance digestive health, improve nutrient absorption, and boost immune responses, ultimately leading to improved overall performance and carcass traits (Didari et al., 2014; El-Sawah et al., 2020; Jeni et al., 2021). Therefore, the objective of the current study was to investigate the effects of different autochthonous LAB as probiotic additives for Muscovy ducks in housing on performance, carcass traits, and serum biochemical parameters.

MATERIALS AND METHODS

The current experiment was conducted at the Faculty of Agrarian Sciences of the Federal University of Amazonas (UFAM), located in Manaus, Amazonas State, Brazil. All experimental procedures were conducted in accordance with the guidelines of the Local Experimental Animal Care Committee, and were approved by the UFAM ethics committee (protocol number 003/2021).

Identification of autochthonous lactic acid bacteria and preparation of probiotics

The intestinal content of 12 domestic Muscovy ducks was used for the isolation of LAB. The content was serially diluted to 10^-5 in Phosphate Buffered Saline (PBS, pH 7.2), plated on MRS agar, and incubated for 24 hours at 37 °C. Selected colonies were inoculated into MRS broth and incubated with agitation at 150 rpm for 24 hours at 37 °C. Randomly selected colonies were purified in MRS broth and validated by Gram staining and catalase test. For further examinations, only Gram-positive and catalase-negative strains were chosen. The isolates were preserved at -20 °C in MRS broth supplemented with 20% (w/v) glycerol.

Genomic DNA was extracted using the PureLink Genomic DNA extraction kit from Invitrogen (Thermo Fisher Scientific^©, Waltham, USA) and the 16S rDNA gene was amplified using the primers: Forward: 5’ CCA TCT CAT CCC TGC GTG TCT CCG ACT CAG ACA CGT AGT ATA CTC CTA CGG RAG GCA GCA 3’; Reverse: 5’ CCT ATC CCC TGT GTG CCT TGG CAG TCT CAG GGA CTA CCA GGG TAT CTA AT 3’. The Asparagin tool available on the EMBRAPA Portal was used to analyze the sequence quality, using the Phred software.

The RDP release database was utilized to deposit data on Bacteria and Archaea, facilitating comparisons with the data of interest to determine bacterial origin, identity percentage, data reliability, and microbiological classification at the genus and species levels (Cole et al., 2009). To validate information from the RDP database, BLAST (Basic Local Alignment Search Tool) alignment was conducted to identify similar regions based on the highest identity percentage, confirming the classification obtained from the initial software (Altschul et al., 1990). Taxonomic classification was performed using bioinformatics tools through a pipeline designed for microbial analyses. Microorganisms were classified using 16S rDNA gene fragment alignment tools, with a similarity index of 97% for species and 95% for genus approximation.

Three strains with promising probiotic potential were identified, classified, and isolated, namely: Enterococcus lactis (BPCD109; Genbank access MF097970.1), Enterococcus ratti (BPCI114; Genbank access MN416973.1), and Enterococcus faecium (BPCJ174; Genbank access MN422615.1). Each LAB strain was individually prepared by reactivating them in MRS broth and incubating at 37°C with agitation at 150 rpm for 24 hours. For biomass production, the bacterial broth was centrifuged at 4,000 rpm for 15 minutes, and the resulting pellets were washed twice with sterile PBS solution. After washing, the pellets from each strain were resuspended in sterile PBS and adjusted to a concentration of 1.5x10⁸ using a spectrophotometer. LAB concentration was confirmed using a 0.5 McFarland standard solution based on turbidity levels (Yosi et al., 2020).

Facilities, animals, diets and experimental design

The aviary used had a ceiling height of 3.25 m, with structural adaptations to improve bird welfare. The temperature and relative humidity were monitored using a digital thermohygrometer, registering averages of 28.3 ºC and 62%, respectively. Throughout the entire experimental period, the Muscovy ducks were monitored for signs of thermal stress caused by the environment, which were observed at no point throughout the study.

Male Muscovy ducks (n = 96) of the creole strain (Rufino et al., 2017a) from the Experimental Farm of the Federal University of Amazonas (Manaus, Brazil) were used. Initially, the day-old male Muscovy ducks (weight = 68.64±1.23 g) were received and housed until 7 days of age in a protective circle, and the floor was covered by wood shavings, tray-type feeders, and cup-type drinkers at a rate of 50 birds/m², with an electric heating source in the center. At 8 days of age (weight = 158.56±2.17 g), the Muscovy ducks were separated into the treatment pens, each with 4 m², tubular-type feeders, hanging-type drinkers, and the floor covered with wood shavings. Experimental management stages were classified as starter (8 to 35 days), grower (36 to 70 days), and finisher (71 to 90 days), during which the birds received feed and water ad libitum. A lighting program similar to that used for slow-growing broilers (20 hours light + 4 hours dark) was used (Wu et al., 2022).

Experimental diets (Table 1) were calculated according to the reference values provided by Rostagno et al. (2024), except for energy and protein (Rufino et al., 2017b), calcium (Feijó et al., 2016), available phosphorus (Costa et al., 2019) and sodium (Santos et al., 2020), which followed the proper requirements of Muscovy ducks. The composition of these diets was analyzed to confirm the proximate composition of the diets according to the methods described by the Van Soest et al. (1991) and AOAC (2019).

Table 1
Experimental diets composition.

Feedstuffs	Quantities (g/kg)
	Starter	Grower	Finisher
Corn 7.88%	577.689	664.061	698.656
Soybean meal 46%	362.806	280.665	241.492
Limestone	10.679	11.111	11.533
Dicalcium phosphate	26.244	24.238	21.889
Sodium chloride	8.342	7.081	5.824
DL-methionine 99%	0.711	1.286	1.266
Vitamin and mineral supplement	5.0001	5.0002	5.0003
Soybean oil	8.529	6.558	14.340
Total (g)	1000.000	1000.000	1000.000
Nutritional levels4
Metabolizable energy, kcal/kg	2,900.00	3,000.00	3,100.00
Dry matter, %	88.134	88.005	88.026
Crude protein, %	21.500	18.000	16.500
Total methionine + cystine, %	0.744	0.705	0.665
Total methionine, %	0.404	0.402	0.382
Total lysine, %	1.153	0.946	0.846
Total threonine, %	0.836	0.718	0.660
Total tryptophan, %	0.270	0.222	0.198
Crude fat, %	3.343	3.334	4.170
NDF, %	11.737	11.555	11.393
ADF, %	4.802	4.457	4.270
Total minerals, %	7.696	7.084	6.601
Calcium, %	1.194	1.137	1.050
Available phosphorus, %	0.600	0.550	0.500
Sodium, %	0.350	0.300	0.250

• ¹
  Vitamin mineral supplement - starter - content in 1 kg of diet = Folic Acid 4 mg, Pantothenic Acid 62.500 mg, Antioxidant 0.0025 g, Biotin 0.2 mg, Niacin 168 mg, Selenium 1.5 mg, Vit. A 33.500 UI, Vit. B1 8.75 mg, Vit. B12 48 mcg, Vit. B2 24 mg, Vit. B6 12.5 mg, Vit. D3 8,000 UI, Vit. E 70 mg, Vit. K3 7.2 mg, Manganese 375 mg, Zinc 250 mg, Iron 250 mg, Copper 40 mg, Iodine 3.75 mg..
• ²
  Vitamin mineral supplement - grower - content in 1 kg of diet = Folic Acid 3.25 mg, Pantothenic Acid 52 mg, Antioxidant 0.0025 g, Niacin 140 mg, Selenium 1.5 mg, Vit. A 28,000 UI, Vit. B1 0.00275 mg, Vit. B12 40 mcg, Vit. B2 20 mg; Vit. B6 10.4 mg, Vit. D3 6,000 UI, Vit. E 50 mg, Vit. K3 6 mg, Manganese 375 mg, Zinc 250 mg, Iron 250 mg, Copper 40 mg, Iodine 3.75 mg.
• ³
  Vitamin mineral supplement - finisher - content in 1 kg of diet = Pantothenic Acid 35.35 mg, Antioxidant 0.0025 g, Niacin 102 mg, Selenium 1 mg, Vit. A 9,800 UI, Vit. B12 23.5 mcg, Vit. B2 12 mg, Vit. D3 2,750 UI, Vit. E 50 mg, Vit. K3 2.75 mg, Manganese 375 mg, Zinc 250 mg, Iron 250 mg, Copper 40 mg, Iodine 3.75 mg.
• ⁴
  Analyzed and calculated levels in Dry Matter.

The experimental design was completely randomized, whereby treatments comprised a control (without use probiotic) and three strains of autochthonous LAB as probiotics administered to the birds (Enterococcus lactis, Enterococcus ratti and Enterococcus faecium), with three replicates of 8 birds each. The probiotic bacteria were administered to the birds orally, according to the methodology described by Olnood et al. (2015), using an automatic pipettor with sterilized 1000 µl tips. The birds received 1 mL/bird of the probiotics (concentration of 1.5 x 10⁸ CFU/mL of bacteria in the provided liquid) daily from day 8 to day 15 of life. Afterward, the same concentration was administered to each bird once a week until they reached 70 days of the experimental period.

Performance and carcass traits

The performance of Muscovy ducks was assessed both within each designated stage and overall. Variables evaluated included feed intake (kg/bird), weight gain (kg/bird), and feed conversion ratio (kg/kg). Feed intake for each stage was calculated by dividing the total feed consumed by the number of birds in each pen. Weight gain was determined by dividing the total pen weight relative to the previous weighing by the number of birds. Feed conversion ratio was computed as the ratio of total feed consumed to weight gain. Combining data from all stages allowed for a comprehensive assessment of these parameters.

At 90 days of age, following a 12-hour fast, all birds from each treatment were weighed, electrically stunned (40 V; 50 Hz), and slaughtered by cutting the jugular vein. Carcasses were then scalded in hot water (60 °C/140 °F for 62 s), plucked, and eviscerated, following the recommendations of Mendes & Patricio (2004) to determine carcass yield (%), and the yields of the main economically significant viscera (liver (g/bird), gizzard (g/bird) and heart (g/bird)) and foot (g/bird)). Commercial cuts (neck, breast, wing, back, thigh, and drumstick) were separated and weighed using an analytical balance (0.01 g), calculating their yields as a percentage relative to the clean carcass.

Serum biochemical parameters

Before slaughter, the Muscovy ducks underwent blood collection. One milliliter of blood was collected from the broilers directly from the ulnar vein, using disposable syringes containing heparin anticoagulant (5000 IU per sample). These samples were immediately centrifuged at 7,000 rpm for 10 minutes to separate the red blood cells for evaluation of hematological parameters from the plasma used for analysis of serum biochemical parameters. These samples were identified and preserved at -4 °C (24.8 °F) throughout the process to be sent to the laboratory. In the analysis of serum biochemical parameters, the plasma samples remaining after centrifugation were subjected to commercial enzymatic-colorimetric assay kits according to the manufacturer’s specific recommendations, and the readings were taken on a mass spectrophotometer (model K37-UVVIS, Kasvi^©, São José dos Pinhais, Brazil) at a specific wavelength for each assay. The biochemical parameters analyzed were the concentrations of total proteins, triglycerides, glucose, cholesterol, and albumin.

Statistical analyses

The data were initially assessed for normality, and necessary transformations were applied. Subsequently, a one-way ANOVA was conducted using the R software (version 4.1.3), following Logan’s (2010) guidelines. Tukey’s honestly significant difference test was used to test the significant differences among the mean values. The results are presented as means and the significant level for differences was set as p<0.05.

RESULTS AND DISCUSSION

Firstly, it is important to highlight that the Enterococcus genus found in this study has a significant capacity to colonize the digestive tract and multiply, showing viable characteristics across a wide range of temperature and pH, and being able to grow in the presence of 6.5% NaCl and 40% bile salts (Willis et al., 2008; Olnood et al., 2015; Qiu et al., 2022), consistent with our in vitro results. In this study, nine strains of LAB were isolated and identified, and the three that stood out based on the Mahalanobis scale were used for tests with the birds, namely: Enterococcus lactis, Enterococcus ratti, and Enterococcus faecium. These strains showed higher sequence identity through BLAST analysis and greater sequence homology through phylogenetic and 16S rDNA sequence homology analysis (Tuli et al., 2014).

Regarding the performance data of the Muscovy ducks subjected to probiotics during the starter stage (8 to 35 days) (Table 2), it was found that there was no significant difference (p>0.05) in feed intake. These results disagree with those found by Willis & Reid (2008) and Alkhalf et al. (2010), who observed a significant difference in feed intake and feed conversion ratio in broilers receiving a diet supplemented with probiotics, even though they used a different bird species. In this context, it is important to emphasize that probiotics do not act as nutrients, but rather as additives that impact the microbial population of their host, especially improving the beneficial microbial population (Gaggia et al., 2010; Olnood et al., 2015). According to Thomas et al. (2015), this may cause positive effects on performance and other benefits depending on the secondary effects that probiotics can provide to the birds.

Table 2
Performance of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives in the starter stage (8 to 35 days).¹

Variables	Probiotics				p-value2	CV3, %
	Control	E. lactis	E. ratti	E. faecium
Feed intake, kg/bird	2.23	2.40	2.29	2.48	0.77	12.07
Weight gain, kg/bird	1.01	0.98	0.98	0.95	0.93	9.88
Feed conversion ratio, kg/kg	2.21c	2.45b	2.34b	2.61a	0.05	11.28

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05)
• ³
  CV = Coefficient of variation.

Weight gain also showed no significant effect (p>0.05) for Muscovy ducks in the starter phase. These results differ from those found by Hassan & Komilus (2020), who, when using 250 mg and 500 mg of lyophilized multi-strain probiotics per kilogram in the starter stage, observed a significant difference (p<0.05) in duckling weight gain, with higher probiotic inclusion resulting in heavier animals. Gaggia et al. (2010) also mentioned that the continuous inclusion of beneficial microorganisms with probiotic potential can induce better functional capacity of the gastrointestinal barrier, which can lead to better nutrient utilization and, consequently, heavier animals (Khattab et al., 2021).

On the other hand, feed conversion ratio showed a significant effect (p<0.05), whereby the control treatment showed the lowest result, meaning that Muscovy ducks subjected to probiotic treatments had a poorer feed conversion ratio. Didari et al. (2014) and Fernandes et al. (2017) suggest that the administration of probiotics, besides naturally causing changes in the composition and activity of intestinal bacteria, may impact the efficiency of digestion and nutrient absorption from the diet, leading to a poorer feed conversion ratio in situations where the birds may not utilize the ingested nutrients as well, or increase their energy expenditure to maintain intestinal health and other physiological processes, which can result in an increased feed intake to maintain the same level of production. In addition, the authors suggested age to be an important factor related to the scenario described above, which may also be linked to the results of this study, where young birds may not be affected or may not initially exhibit the beneficial effects of probiotic administration. This indicates that probiotics should be administered long-term to fully express their effects.

Regarding the performance of Muscovy ducks during the grower stage (Table 3), no significant effects (p>0.05) were observed on feed intake and weight gain from using bacteria as probiotics. These findings differ from Abou-Elnaga et al. (2018), who observed improved parameters in ducks after 70 days using a commercial probiotic based on Bacillus subtilis and Bacillus licheniformis. In terms of feed conversion ratio, both the control group and Muscovy ducks treated with Enterococcus lactis showed lower results (p<0.05), indicating better feed conversion ratio for the Muscovy ducks treated with this probiotic strain. This suggests enhanced digestion and nutrient absorption at the intestinal level with the presence of this strain (Hassan & Komilus, 2020).

Table 3
Performance of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives in the grower stage (36 to 70 days).¹

Variables	Probiotics				p-value 2	CV3, %
	Control	E. lactis	E. ratti	E. faecium
Feed intake, kg/bird	3.69	3.66	3.63	3.74	0.94	5.54
Weight gain, kg/bird	1.09	1.08	0.98	1.06	0.90	16.47
Feed conversion ratio, kg/kg	3.39c	3.39c	3.70a	3.53b	0.05	13.27

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05)
• ³
  CV = Coefficient of variation.

Regarding the performance of Muscovy ducks during the finisher stage (Table 4), significant effects (p<0.05) were observed when using LAB as probiotics on all variables analyzed. Muscovy ducks in the control group exhibited higher weight gain and better feed conversion ratio compared to those subjected to LAB as probiotics, with even the Muscovy ducks subjected to Enterococcus faecium presenting a lower feed intake. Previous studies suggest that the gastrointestinal tract of birds, particularly non-chicken species, may not support the survival of LAB under certain environmental conditions (Gaggia et al., 2010; Didari et al., 2014; Jeni et al., 2021; Khattab et al., 2021). While LAB have been effective probiotic additives for chickens (Alkhalf et al., 2010; Gaggia et al., 2010; Hassan & Komilus, 2020), the impact of LAB on the microbial population in the Muscovy duck’s intestinal region might be limited due to specific environmental conditions that differ these from other poultry species (Abou-Elnaga et al., 2018; El-Sawah et al., 2020).

Table 4
Performance of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives in the finisher stage (71 to 90 days).¹

Variables	Probiotics				p-value 2	CV3, %
	Control	E. lactis	E. ratti	E. faecium
Feed intake, kg/bird	2.71b	2.83a	2.76b	2.55c	0.05	6.84
Weight gain, kg/bird	1.00a	0.74b	0.78b	0.71b	0.05	2.44
Feed conversion ratio, kg/kg	2.71c	3.82a	3.54b	3.59b	0.05	2.55

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05)
• ³
  CV = Coefficient of variation.

No significant effect (p>0.05) was observed from using LAB as probiotics on the general feed intake of the Muscovy ducks when evaluating the results of the general performance of Muscovy ducks (Table 5), as well as that of the separated stages. However, Muscovy ducks in the control group exhibited higher (p<0.05) weight gain and better (p<0.05) feed conversion ratio compared to those subjected to LAB as probiotics. These results contrast with findings from Lokapirnasari et al. (2022), who reported improvements in weight gain among Indonesian ducks treated with Lactobacillus casei, Lactobacillus lactis, and Bifidobacterium spp. in conjunction with Moringa oleifera extract. Similarly, Khattab et al. (2021) observed no significant performance differences when Pekin ducks were supplemented with Lactobacillus acidophilus. Such discrepancies highlight the complex interactions between probiotic strains and specific avian species, suggesting that the efficacy of probiotics may vary based on environmental conditions, genetic factors, and overall diet quality.

Table 5
General performance of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives.¹

Variables	Probiotics				p-value 2	CV3, %
	Control	E. lactis	E. ratti	E. faecium
Feed intake, kg/bird	8.65	8.90	8.69	8.77	0.71	2.99
Weight gain, kg/bird	3.10a	2.81b	2.75b	2.72b	0.05	11.77
Feed conversion ratio, kg/kg	2.80b	3.17a	3.16a	3.22a	0.05	13.40

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05)
• ³
  CV = Coefficient of variation.

Moreover, the literature indicates that nutrient utilization in Muscovy ducks is influenced by a multitude of factors, including diet digestibility and nutritional composition (Hassan & Komilus, 2020; Arias-Sosa & Rojas, 2021; Lokapirnasari et al., 2022). This multifactorial nature underscores the need for a comprehensive understanding of how different probiotic strains interact with the gut microbiota and affect physiological processes (Lokapirnasari et al., 2022; Maquiné et al., 2024). In this context, even environmental stressors such as temperature and housing conditions could also play a role in how effectively probiotics contribute to performance outcomes (Khattab et al., 2021; Maquiné et al., 2024).

Intentional inoculation of avian microbiota with probiotic bacteria aims to enhance beneficial microbial populations and deter pathogenic enteric microorganisms (Oliveira et al., 2014). Successful colonization in the gastrointestinal tract, particularly in early development stages, depends on the specific microorganism used and its impact on the avian intestinal microbiota. This can range from a positive influence to no significant effect, maintaining nutrient use and performance at stable levels (Khattab et al., 2021). Therefore, while LAB showed potential in some studies (Alkhalf et al., 2010; Thomas et al., 2015; Yosi et al., 2020), their application in Muscovy ducks did not yield the anticipated benefits, pointing to the necessity for further research into strain-specific responses and the development of tailored probiotic strategies that align with the unique needs of different poultry species.

In carcass traits analysis (Table 6), Muscovy ducks in the control group had higher slaughter weights (p<0.05) and greater yields of neck and drumstick (p<0.05) compared to those given LAB as probiotics. However, Muscovy ducks treated with Enterococcus lactis and Enterococcus ratti showed similar slaughter weights to the control group and had higher back yield (p<0.05). These results are consistent with studies by Alkhalf et al. (2010) and Thomas et al. (2015), which found beneficial effects of probiotics on birds’ slaughter weight. Combining probiotic bacteria with dietary ingredients can enhance nutrient viability during intestinal transit (Yosi et al., 2020) and improve digestive enzyme activity and nutrient absorption in the gastrointestinal tract, potentially leading to better overall nutrient utilization (Park et al., 2016).

Table 6
Carcass traits of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives.¹

Analysis	Variables	Probiotics				p-value 2	CV3, %
		Control	E. lactis	E. ratti	E. faecium
Carcass yield and viscera	Slaughter weight, kg/bird	3.01a	2.98ab	2.99ab	2.81b	0.03	6.09
	Carcass yield, %	87.40b	87.96b	88.02b	91.15a	0.03	4.62
	Foot, g/bird	91.33	91.66	91.33	87.66	0.43	5.22
	Liver, g/bird	38.66	41.33	40.66	38.33	0.72	13.12
	Gizzard, g/bird	73.66b	80.66a	75.00b	72.66b	0.05	8.68
	Heart, g/bird	22.33	21.33	22.33	20.33	0.52	12.21
	Pancreas, g/bird	2.70b	3.13a	2.76b	2.66b	0.05	5.24
Commercial cuts	Neck, %	11.21a	10.49b	9.19c	8.91c	0.03	13.79
	Wings, %	16.31	16.61	17.03	16.67	0.71	6.21
	Drumstick, %	13.07a	12.66b	9.84c	11.59b	<0.01	14.96
	Thigh, %	9.20	8.91	10.74	9.27	0.23	17.62
	Breast, %	24.72c	24.67c	25.42b	27.49a	0.04	13.16
	Back, %	25.49c	26.66b	27.78a	26.07b	0.05	8.81

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05).
• ³
  CV = Coefficient of variation.

In the carcass yield results, it was observed that Muscovy ducks subjected to Enterococcus faecium presented higher (p<0.05) carcass and breast yields. Additionally, there were gradual increases for this variable compared to the control group as the birds were subjected to the probiotics, with the carcass results of all probiotic treatments being numerically higher than the control. These results align with those reported by Abou-Elnaga et al. (2018) and contrast with those reported by Hassan & Komilus (2020), suggesting that the inclusion of probiotics may positively influence carcass yield and composition, potentially due to improved nutrient absorption and gut health. The variations in findings could be attributed to differences in the probiotic strains used, the duration of supplementation, or specific management practices in each study.

The fact that treatments that received probiotics achieved higher carcass yield compared to the control group may indicate that the administered LAB provided a positive result regarding the formation of the Muscovy ducks’ carcass structures through better nutrient availability (Gaggia et al., 2010). In this context, Ardiansah et al. (2020) found a significant effect on carcass yield (p<0.05) when providing powder and encapsulated probiotic to local Indonesian ducks, stating that probiotic supplementation may have contributed to increasing protein availability and consequently provided a better carcass yield.

The results also demonstrated that using bacteria as probiotics significantly influenced (p<0.05) the weight of the gizzard and pancreas, indicating that the use of Enterococcus lactis stimulated the growth of these internal organs, which differs from the results found by Ardiansah et al. (2020), who did not obtain significant results for the same organs. According to Tuli et al. (2014), there may be a correlation between the use of a LAB as probiotics and the increase of certain organs, especially those related to the digestive system, with the administration of LAB stimulating metabolic pathways through greater nutrient availability, which can cause higher activity of these organs and consequently stimulate their growth.

In the results of serum biochemical parameters of Muscovy ducks in housing (Table 7), significant effects (p>0.05) were observed from using LAB as probiotics on all variables analyzed. Muscovy ducks from the control group showed higher (p<0.05) total protein concentration values, while the groups supplemented with LAB as probiotics had reduced and similar values among them, ranging from 0.31 to 0.32 g/dL. This suggests that supplementation with LAB may have reduced total protein levels compared to the control group, which may be related to their own properties, which include enhancing gut microbiota balance and improving the intestinal barrier function (Haghighi et al., 2005; Gaggia et al., 2010). By doing so, they can facilitate better nutrient absorption and utilization, potentially leading to more efficient protein metabolism and resulting in lower circulating levels of total protein, as the body may then utilize proteins more effectively for physiological processes rather than allowing them to accumulate in the bloodstream (Gaggia et al., 2010).

Table 7
Serum biochemical parameters of Muscovy ducks in housing supplemented with autochthonous lactic acid bacteria as probiotic additives.¹

Variables	Probiotics			p-value 2	CV3, %
	Control	E. lactis	E. ratti	E. faecium
Total proteins, g/dL	0.46a	0.31b	0.32b	0.32b	0.05	6.22
Triglycerides, mg/dL	71.25b	59.07c	74.71b	100.87a	0.05	5.23
Glucose, mg/dL	165.83b	158.95c	163.61b	174.90a	0.05	7.15
Cholesterol, mg/dL	124.87b	109.47c	101.32c	149.60a	0.05	3.07
Albumin, mg/dL	2.48b	2.51b	1.82c	3.11a	<0.01	2.46

• ¹
  All data represent the mean of 24 replicates (Muscovy ducks) per treatment.
• ²
  Means followed by lowercase letters in lines differ by Tukey’s test (p<0.05).
• ³
  CV = Coefficient of variation.

On the other hand, Muscovy ducks subjected to Enterococcus faecium presented higher (p<0,05) concentrations of triglycerides, glucose, cholesterol and albumin, suggesting that Enterococcus faecium supplementation may have distinct metabolic effects on these serum biochemical parameters in Muscovy ducks. Enterococcus faecium is a type of LAB commonly used as a probiotic in animal nutrition (El-Sawah et al., 2020). While LAB are generally recognized for their beneficial effects on gut health and overall well-being, the observed metabolic changes in the control group raise questions about the specific interactions between Enterococcus faecium and metabolic pathways in Muscovy ducks (Alagawany et al., 2018; El-Sawah et al., 2020).

One potential explanation for the increased triglycerides, glucose, cholesterol, and albumin levels could be alterations in nutrient absorption, metabolism, or utilization induced by Enterococcus faecium supplementation. Probiotics can influence the gut microbiota composition and activity, which in turn may impact nutrient absorption and metabolism (Butel, 2014; El-Sawah et al., 2020). Changes in gut microbiota composition could potentially lead to increased absorption of triglycerides, glucose, and cholesterol from the diet, contributing to elevated serum levels of these compounds (Didari et al., 2014; Alagawany et al., 2018).

CONCLUSIONS

The use of autochthonous LAB as probiotics in Muscovy duck diets had varying effects across growth stages, indicating complex interactions with avian physiology. While no significant impacts were observed on feed intake and weight gain during the starter stage, feed conversion ratio was compromised. A similar variability was observed during the grower and finisher stages, whereby probiotics resulted in lower performance results compared to the control group. Enterococcus lactis showed promise in improving feed conversion ratio and carcass traits, while Enterococcus faecium affected all serum biochemical parameters. These results suggest that the probiotics’ effectiveness may depend on the Muscovy ducks’ developmental stage and the specific probiotic strains used. These findings highlight that future research can investigate the mechanisms by which these strains influence other variables beyond feed conversion ratio and serum biochemical parameters, as well as explore the application of probiotics under diverse management conditions to validate the consistency of the effects observed in this study. Comparative studies with other bird species would also be valuable to understand the broader applicability of the identified probiotics.

ACKNOWLEDGMENTS

To Fundação Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) (Grant No. 2448/2022; Process No. 88881.692376/2022-01; PDPG-POSDOC-Programa de Desenvolvimento da Pós-Graduação (PDPG) Pós-Doutorado Estratégico), the Fundação de Amparo à Pesquisa do Estado do Amazonas (FAPEAM), and the Programa de Pós-Graduação em Biotecnologia (PPGBIOTEC) of the Instituto de Ciências Biológicas (ICB) at the Universidade Federal do Amazonas (UFAM) for their support in the development of this study.

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• Funding
  None.
• Data availability statement
  The data of this study are available from the corresponding author upon reasonable request.

Edited by

• Section editor:
  Irenilza A. Nääs

Data availability

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

Publication Dates

• Publication in this collection
  25 Nov 2024
• Date of issue
  2024

History

• Received
  24 Apr 2024
• Accepted
  08 Sept 2024