Research Article

Bioactive Substances and the Effect of Dietary Dayak Onion Meal (Eleutherine bulbosa) Supplementation on Performance and Intestinal Health of Superior Native Chicken

Azhar Syafiq Imanullah1, Nanung Danar Dono2, Bambang Ariyadi2, Aan Andri Yano3*

1Faculty of Agriculture, Universitas Lambung Mangkurat, Jl. A. Yani, Banjarbaru, South Kalimantan 70714, Indonesia; 2Faculty of Animal Science, Universitas Gadjah Mada, Jl. Fauna No. 3 Bulaksumur, Yogyakarta 55281, Indonesia; 3Vocational School, Universitas Sebelas Maret, Jl. Ir. Sutami No.36, Surakarta, Central Java 57126, Indonesia.

Received | June 11, 2025; Accepted | July 20, 2025; Published | August 16, 2025

*Correspondence | Aan Andri Yano, Vocational School, Universitas Sebelas Maret, Jl. Ir. Sutami No.36, Surakarta, Central Java 57126, Indonesia; Email: [email protected]

Citation | Imanullah AS, Dono ND, Ariyadi B, Yano AA (2025). Bioactive substances and the effect of dietary dayak onion meal (Eleutherine bulbosa) supplementation on performance and intestinal health of superior native chicken. Adv. Anim. Vet. Sci., 13(9):1870-1879.

DOI | https://dx.doi.org/10.17582/journal.aavs/2025/13.9.1870.1879

ISSN (Online) | 2307-8316

Copyright: 2025 by the authors. Licensee ResearchersLinks Ltd, England, UK.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

INTRODUCTION

The demand for native chicken products in Indonesia has been steadily rising (Hidayat and Asmarasari, 2015). However, production has not kept pace, resulting in a supply deficit. To address this issue, Kampung Unggul Badan Litbang Pertanian (KUB), or Superior Native Chicken, has been developed as a viable alternative. KUB chickens were introduced by the Ciawi Bogor Animal Research Institute under the Indonesian Agency for Agricultural Research and Development (IAARD). These chickens were selectively bred for improved egg production and reduced broodiness while maintaining the genetic characteristics of Indonesian native chickens (Muslim et al., 2021). Rahayu et al. (2021) stated that KUB chickens exhibit significantly higher egg production than ordinary native chickens. Additionally, they demonstrate strong adaptability to harsh environmental conditions, which enhances their suitability for smallholder farming systems in Indonesia.

Although KUB chickens offer promising productivity, they are still susceptible to infections by harmful pathogenic microbes. Poultry production in Indonesia used to rely heavily on antibiotics to tackle pathogenic microbial infections. Antibiotics have been widely used as growth promoters to enhance performance (Miles et al., 2006; Ronquillo and Hernandez, 2017) with the main mechanism as antibacterial which is useful in inhibiting or killing pathogenic bacteria in the small intestine, thereby improving nutrient absorption. Mehdi et al. (2018) added that sub-therapeutic doses of antibiotics were historically administered to broiler chickens to accelerate growth, improve health, and enhance productivity. Common antibiotics added to poultry feed include penicillin, tylosin, virginiamycin, and zinc-bacitracin.

The continuous use of antibiotics in animal feed has raised significant concerns regarding the emergence of antibiotic-resistant bacteria and the accumulation of antibiotic residues in livestock products (Xu et al., 2022). To mitigate these risks, the Government of Indonesia, along with several other countries, has implemented a ban on the use of antibiotics as Antibiotic Growth Promoters (AGPs) (Hibbard et al., 2023). In Indonesia, this ban is regulated under the Minister of Agriculture Regulation Number 14 of 2017 concerning the Classification of Veterinary Drugs. This policy represents a strategic step to control antimicrobial resistance and protect consumers from exposure to antibiotic residues in livestock products (Oladeji et al., 2025).

However, the ban on the use of antibiotics has inadvertently led to challenges in poultry production, particularly the resurgence of pathogenic microorganisms (Haque et al., 2020). Digestive tract diseases remain a persistent issue, as the use of AGPs can trigger an increase in antibiotic resistance in pathogenic bacteria. This resistance may worsen infections and contaminate animal products, ultimately posing a threat to public health (Obianwuna et al., 2024). One of the most prevalent and challenging diseases is Necrotic Enteritis (NE), which primarily affects chickens older than four weeks. NE is characterized by rapid onset and sporadic bacterial attacks, particularly the jejunum and ileum, causing severe necrosis of the small intestine mucosa (Gautam et al., 2024). This disease poses a substantial threat to poultry health and productivity.

To address these challenges, various alternatives to antibiotics have been found, one of which is phytobiotics. Phytobiotics are feed additives that are purely derived from plants added to animal feed to improve production performance and livestock health (Alghirani et al., 2021). Phytobiotics are recognized for their efficacy because they have bioactive compounds with multiple beneficial properties, such as antibacterial, antioxidant, and anti-inflammatory activities (Gheisar et al., 2018). These compounds can be derived from various plant parts, including leaves, stems, and roots (Valli and Kavitha, 2023). One promising plant that may serve as an alternative to antibiotics is Dayak onion.

The Dayak onion (Eleutherine bulbosa) is a plant with significant potential as a natural source of antioxidants (Sekarsari et al., 2024). Its phytochemical content includes alkaloids, tannins, glycosides, flavonoids, and phenolics, that are able to exhibit antioxidant properties (Mukti et al., 2023). Many studies highlight the efficacy of Dayak onion in livestock and the animal products. Riadh et al. (2024) demonstrated its role in maintaining chicken meat pH for up to 9 hours, while Monasdir et al. (2024) found that Dayak onions up to 4% inclusion significantly affect pH, water content, protein content, antioxidants and texture profile in chicken sausages. Additionally, the combination of Dayak onion extract and Lactobacillus acidophilus at 0.3% significantly enhances meat characteristics and growth performance through the increase in antioxidant activity of broiler chickens (Yuanita et al., 2019). Moreover, concentration of 12.5-15.0 mg/mL of Dayak onion extract effectively reduce total E. coli and S. aureus while maintaining chicken meat quality under refrigeration (Romadhon et al., 2023). However, some studies indicate limitations. Haerul et al. (2024) found that Dayak onion meal had no significant effect on quail performance and egg physical quality. Similarly, the level of Dayak onion extracts up to 20 g in the drinking water does not significantly affect the body weight gain, feed consumption, and water consumption of broiler chicken (Zakiyatulyaqin et al., 2020).

These characteristics make the Dayak onion a promising candidate for further research into its potential as a feed additive and an alternative to antibiotics in poultry nutrition. Its reported antimicrobial, antioxidant, and meat quality-enhancing properties (Yuanita et al., 2019; Romadhon et al., 2023; Monasdir et al., 2024), suggest that it could play a significant role in improving livestock productivity and product safety. However, despite these promising findings, research on its application in native chicken breeds, particularly KUB chicken, remains limited. Most existing studies have focused on broilers, quails, or meat preservation, leaving a gap in understanding its effect on indigenous poultry breeds.

Therefore, this study aimed to evaluate the bioactive substance content of Dayak Onion Meal (DOM) and assess its effects on the performance and intestinal health of superior native chickens. By investigating its influence on performance, gut microbiota, and intestinal health, this research will provide valuable insights into the potential of DOM as a functional feed additive. The findings could contribute to the development of natural growth promoters in poultry nutrition, offering a sustainable and antibiotic-free alternative for improving livestock productivity while supporting gut health in native chicken breeds.

MATERIALS AND METHODS

Time and place of study

The experiment was conducted from June to September 2024. The chickens were sourced from and reared at the experimental farm of the Parent Seed Garden of the Agricultural Technology Assessment Institute, Agency for Agricultural Research and Development, South Kalimantan, Indonesia. Meanwhile, microbial and chemical analyses were performed at Universitas Gadjah Mada, Indonesia. Additionally, morphological data collection was carried out at the Laboratory of Pathology, Banjarbaru Veterinary Center, South Kalimantan, Indonesia. All animal use protocols for the experimental research were reviewed and approved by a veterinarian who followed the ethical guidelines outlined in the Law of Republic of Indonesia number 18, 2009 on animal husbandry and health.

DOM making

Fresh Dayak onion tubers were obtained from a commercial shop in Hulu Sungai Tengah Regency, South Kalimantan Province. The tubers were then dried in an oven (Memmert Incubator Oven INB200, Memmert, Schwabach, Germany) at 60 °C for approximately 2 days. The dried tubers were then ground into a fine powder using a Wiley mill (Model 4 Wiley Mill, Thomas Scientific, USA) equipped with a 1-millimeter steel screen.

Research design

A total of 120 one-day-old KUB native chickens were randomly allocated to 5 treatment groups, each with 4 replicates of 6 birds. Since Indonesian native chickens are slow-growing, and their digestive system is underdeveloped during the starter phase (Tsado et al., 2019), all the birds were fed commercial diet in a mash and crumble form during first 30 days to achieve notable impacts. Commencing from day 31, DOM treatments were administered to the birds until day 90. The experiment applied a completely randomized design with the following treatment groups: T0 (Control)= 0% (basal diet without adding DOM), T1 = Control + 0.5% DOM, T2 = Control + 1.0% DOM, T3 = Control + 1.5% DOM, and T4 = Control + 2.0% DOM. The birds were kept in standard environmental rearing conditions under a colony system. Rice hulls were used as a bedding material in a floor cage. A light was turned on for 12 hours during the night. The feeding was conducted twice daily (morning and evening) according to the following schedule: 50g/chicken/day during week 5 and 6, 65g/chicken/day during week 7 and 8, 75g/chicken/day during week 9 until 11, and 100g/chicken/day during week 12 until harvest. Drinking water was given ad libitum. Composition and nutrient content of basal diet are provided in Table 1.

 

Table 1: Composition and nutrient content of basal diet.

Feedstuff	Proportion (%)
Yellow corn	41.00
Rice bran	17.00
Soybean meal	20.00
Crude palm oil	3.00
Palm kernel meal	17.00
Premix (vitamin and mineral)1	1.50
Top mix2	0.50
Total	100
Chemical composition
Metabolizable Energy (kcal/kg)	2711.24
Crude protein (%)	17.24
Crude fiber (%)	6.22
Crude fat (%)	6.14
Calcium (%)	1.18
Available Phosphorus (%)	0.40
Lysine (%)	0.31
Methionine (%)	0.10

 

Note: 1Premix of vitamins and minerals per kg contains Steam Bone Meal 50%, Calcium 48.72%, Manganese 0.4%, Iodine 0.005%, Ferrum 0.3%, Cuprum 0.02%, Zinc 0.25%, Magnesium 0.29%, Vitamin D3 75,000 IU, Vitamin B12 450 mcg. 2The composition of each mg of Top Mix Mineral-Vitamin contains Calcium 32.5%; Phosphorus 1.0%; Ferrum 6.0 g; Manganese 4 g; Iodine 0.075 g; Zinc 3.75 g; Vitamin B12 0.5 mg; Vitamin A 300,000 IU; Vitamin D3 50,000 IU.

 

Data collection

Phytochemical substances of DOM were analyzed based on modified method of Fitriansyah et al. (2021). Animal performances specifically weight gain was determined by weighing the chickens body weight every weekend using digital weighing scale (JEOL JEM 1400 Pluss, Jeol, Peabody, USA). Meanwhile, feed consumption was calculated by subtracting the remaining feed from the amount initially provided to the chickens in each cage during the 60-day feeding period. The total feed consumed in each cage was then divided by six, corresponding to the number of native chickens in each cage. The feed conversion ratio (FCR) was calculated based on the comparison between the amount of feed consumed and weight gain, which was taken every weekend. The results of the calculation are averaged to obtain the feed conversion during the study.

The method for histomorphological measurement of the small intestine refers to the study by Dono (2012). One sample of native chicken from each replication, representing growth performance, was taken from the middle portion of the jejunum, approximately 6 cm long, with the intestinal contents removed. The sample was prepared for viscosity (consistency) testing of the digesta, and the mucosa was cleaned by spraying with aquades. The jejunum was then cut into three equal sections. The jejunum was preserved using a 10% formalin buffer solution, gently agitated, and stored in tightly closed bottles until further processing. The jejunum samples were then removed from the formalin bottles and dehydrated using a series of increasing alcohol concentrations (70%, 90%, and 100%), cleaned with xylene solution, and embedded in paraffin solution. The jejunum was then sectioned (preparations) were placed on slides for staining with hematoxylin and eosin. The jejunum preparations were observed under a microscope (Olympus Corp., Tokyo, Japan) equipped with a camera (Optilab Advance, Miconos, Indonesia) to capture images and a micrometer to measure the villus height (VH), crypt depth (CD), and the villus-to-crypt depth ratio.

The calculation of intestinal microbiota was performed using the Total Plate Count (TPC) method. The variables observed included the colony count of lactic acid bacteria and Salmonella sp. The digesta was collected and placed into sterile bottles. The TPC media used were Peptone Glucose Yeast Agar for lactic acid bacteria, with a composition similar to de-Man’s, Rogosa, and Sharpe Agar (MRSA), and Salmonella Shigella Agar (SSA) for Salmonella sp. The analysis of lactic acid bacteria population was based on the method of Altaf et al. (2006), while Salmonella sp. was analyzed according to the method of Salehi et al. (2005).

Data analysis

Phytochemical substances of DOM were analyzed descriptively. Data of animal performance, histomorphology, gut microbiota are presented as mean ± standard of deviation and mean ± standard error mean. Data were analyzed using ANOVA test using IBM® SPSS Statistics 25.0 for Windows. Duncan’s new Multiple Range Test (DMRT) was then used to separate means when ANOVA revealed significant effects. This test was used to determine the effect of DOM supplementation on performance, gut microbiota, and intestinal health of KUB native chicken using a significance level of P < 0.05. If the P value is close to 0.01, the result indicates a highly significant effect on these observed variables.

RESULTS AND DISCUSSION

Phytochemical substances of DOM

Phytochemical screening of DOM revealed the presence of several bioactive compounds, with phenols being the most abundant, followed by flavonoids, tannins, saponins, alkaloids, and steroids (Table 2). These results confirm that DOM contains a variety of bioactive substances in notable concentrations. According to Susilawati et al. (2022), Dayak onions contain several bioactive compounds such as alkaloids, steroids, glycosides, flavonoids, phenolics, saponins, and tannins. These results were also confirmed by Supomo et al. (2019), who reported that Dayak onion tubers contain phytochemicals, including triterpenoids, flavonoids, and phenolics. These compounds are known for their natural antioxidants, anti-inflammatory, and antibacterial properties.

 

Table 2: Phytochemical screening result of DOM.

Chemical content in total	Result (% / 100g)
Flavonoid	3.31
Saponin	1.25
Tannin	2.76
Alkaloid	0.66
Phenol	5.56
Steroid	0.58

 

The results of the phytochemical screening for the total flavonoid content of DOM in this study were 3.31% in g/100g. This result is higher compared to the study by Silitonga et al. (2020), which reported a flavonoid content of 0.431% in g/100g in Dayak Onion flour. Additionally, this study found a phenolic content of 5.56% in DOM. These findings are significantly higher than the study by Yuswi (2017), who reported that Dayak Onion powder contained 0.844% phenols. This discrepancy may be attributed to differences in sample preparation, extraction methods, and analytical protocol. Yuswi (2017) used an ultrasonic-assisted extraction with different solvent systems, which may have resulted in lower efficiency. In contrast, this study applied a modified method from Fitriansyah et al. (2021) that directly analyzed DOM powder using a spectrophotometric assay optimized for our sample matrix.

Furthermore, the drying method used in this study (oven drying at 60°C for 2 days) may have influenced the phenol content. While prolonged heating can potentially degrade heat-sensitive phenolic, moderate drying temperatures like 60°C have been shown to preserve phenolic compounds effectively. For instance, oven drying at 60°C provided greatest conservation of phenolics in pomegranate peel flour (Wanderley et al., 2023). Similarly, Babaei et al. (2025) reported oven-drying at 70°C in Capparis spinosa L. fruits resulted in a total phenolic content of 5.3 mg GAE/g, which is comparable to our result in this study.

These findings suggest that the high phenolic content in DOM observed in this study is not anomalous but consistent with data from other plant sources processed at similar temperatures. Moreover, it is plausible that prolonged drying at moderate temperature not only preserved but also enhanced phenolic extractability by breaking down plant cell structures. The antioxidant properties of DOM are therefore likely associated with its high levels of flavonoids and phenolic compounds. Phenolic compounds, characterized by hydroxyl groups attached to aromatic rings, are known to donate hydrogen atoms, allowing DPPH (2,2-diphenyl-1-picrylhydrazyl) radicals to be reduced to a more stable form (Pereira et al., 2009).

This study also identified antinutritional factors (ANFs). ANFs are able to reduce nutrient absorption and digestion, impairing growth performance in poultry (Yano et al., 2024), while it was also reported that combination of tannin and saponin in ruminant feed is able to significantly reduce ammonia production (Jayanegara et al., 2020). In addition, poultry is more sensitive to tannin compared to ruminant (Çalişlar, 2018). Thus, determining ANFs presence is crucial in this study as the objects are native chicken. Here, tannin and saponin are two mains observed ANFs.

Animal performance

Statistical analysis of the effect of DOM supplementation on the growth performance of native chickens, presented in Table 3, showed that the dietary inclusion of DOM did not result in significant changes in any growth performance parameters (p>0.05). It implies that the growth performance of native chickens observed in the study were relatively similar across all treatments including feed intake (FI), feed conversion ratio (FCR), body weight gain (BWG), and final weight. Similar findings are observed in broiler chickens up to 15% in the diet by Silitonga et al. (2020) and up to 20% in drinking water by Zakiyatulyaqin et al. (2020). Given that FI is main factor influencing other animal performances (Abdollahi et al., 2018), these unexpected findings are likely attributed to the low supplementation level, which may have been insufficient to manifest any noticeable impact from the addition of DOM.

Classen (2017) stated that factors influencing FI include nutrient content, livestock age, environment, diseases, and the form and manufacturing process of the feed. In this study, regardless of the treatment, all birds received similar nutrient content, were of the same age, had the same feed form, and were housed in similar facilities. However, no animal diseases were observed in this study. Thus, manufacturing process of the feed appears to be influencing factor for the observed lack of significant difference in FI.

The scordinin content in Dayak onions is speculated to be the cause of the observed results in this study. Scordinin is a volatile compound found in Dayak onion, which makes it prone to evaporation when exposed to heat during the feed heating process (Zakiyatulyaqin et al., 2020). Scordinin serves as a natural growth promoter by stimulating cell growth and repairing damaged body cells (Berliana et al., 2018). However, its effectiveness may have been compromised due to evaporation that occurs during the production of DOM. Additionally, scordinin levels were not measured in this study. Therefore, the extent of its presence in the finished feed and its potential contribution to the observed outcomes remain uncertain.

Histomorphology of jejunal small intestine

The growth of chickens is influenced by the digestion and absorption of nutrients, which are linked to the morphological and functional development of the small intestine (Lisnahan and Nahak, 2020). This study resulted in no significant differences were observed in the height or width of the jejunal villi among treatments (p>0.05), although DOM supplementation significantly influenced crypt depth (p<0.05) and the villus height to crypt depth ratio in the jejunum (p<0.05) (Table 4). To our best knowledge, unfortunately, no single studies have reported the histomorphological effects of DOM. However, we can speculate that the insignificant results for VW and VH

 

Table 3: Effect of DOM supplementation on the chicken performances.

Treatment	FI (g)	FCR	BWG (g)	Final weight (g)
T0	2147.96±20.90	2.51±0.33	688.12±105.19	1119.27±78.97
T1	2183.25±46.41	2.32±0.18	745.31±48.15	1241.37±38.66
T2	2129.32±38.62	2.42±0.20	746.36±67.32	1315.80±271.16
T3	2150.10±17.24	2.24±0.30	766.41±100.91	1429.92±155.69
T4	2139.21±20.69	2.46±0.33	695.38±83.80	1226.59±161.64
P value	0.741	0.651	0.618	0.144

 

Note: T0 (Control) = 0% (basal diet without adding DOM), T1 = Control + 0.5% DOM, T2 = Control + 1.0% DOM, T3 = Control + 1.5% DOM, and T4 = Control + 2.0% DOM.

 

Table 4: Effect of DOM supplementation on histomorphology of jejunal small intestine.

Parameter	T0	T1	T2	T3	T4	SEM	P-value
Villus height (VH) (ɥm)	1082.20	1155.74	962.61	1103.46	987.26	30.313	0.207
Villus width (VW) (ɥm)	262.68	211.32	207.99	252.63	229.81	15.776	0.844
Crypt depth (CD) (ɥm)	396.72bc	594.31a	614.29a	451.85b	330.82c	37.777	0.002
VH:CD ratio	2.73ab	1.94bc	1.56c	2.46ab	3.01a	0.188	0.023

 

Note: Means within a row without a common superscript differ significantly (P < 0.05). T0 (Control) = 0% (basal diet without adding DOM), T1 = Control + 0.5% DOM, T2 = Control + 1.0% DOM, T3 = Control + 1.5% DOM, and T4 = Control + 2.0% DOM.

 

are likely due to the synergistic effect of the negligible FI. Lisnahan and Nahak (2020) reported that the increase in villi size (VH and VW) is positively correlated with feed consumption and growth. It is plausible that the lack of significant growth in VH or VW resulted from insufficient nutrient intake, which in turn could not significantly optimize animal performance. Furthermore, Ravindran and Reza (2021) emphasized that the development of VH and VW plays a significant role in nutrient absorption for chicken. An increase in VW and VH of the small intestine is generally associated with an expansion in its surface area (Kiela and Ghishan, 2016), suggesting that DOM supplementation may not have been significantly enhanced the surface area for nutrient absorption. Additionally, the growth in length and diameter of intestinal segments enhances the surface area available for nutrient uptake.

In contrast, both CD and the VH:CD ratio was affected significantly by the DOM supplementation. For CD, a significant increase was observed at T1 and T2, followed by a gradual, significant decrease at T4. In the case of the VH:CD ratio, however, a significant decrease was noted at T2, followed by a notable increase at T4. Kavoi et al. (2016) explained that in the crypts, cell proliferation occurs through mitosis, and these newly formed cells migrate to the villus tip, where they serve as a protective barrier for the small intestinal cells. The observed increase in T1 and T2 suggest that a 0.5-1.0% DOM supplementation may delay the migration of newly formed epithelial cells to the tip of the villus, potentially impacting the antimicrobial and detoxifying functions of the small intestine. In contrast, the decrease in CD at T4 (2.0% DOM) suggests a potential improvement in the efficiency of cell renewal. Additionally, Marchewka et al. (2021) emphasized that shallow crypts indicate that villi can survive for an extended period without requiring cell renewal. Furthermore, a higher ratio of VH:CD is associated with increased villus area. This aligns with the observations that T4, which had lowest CD value and the highest VH:CD ratio, suggest improved cell turnover and nutrient absorption with higher DOM supplementation.

We speculate that these findings result from the antibacterial activity of DOM, which indirectly supports crypt and villus performance. As presented in Table 2, DOM contains various phytochemical contents. These contents are able to serve as antibacterial, which are driven by their ability to disrupt key cellular processes especially T4. Flavonoids, as the largest group of phenol compound, inhibit bacterial growth by targeting nucleic acid synthesis (Hasnat et al., 2024), alkaloids functions as an anti-microtubule agent, inhibiting mitosis by blocking cell from progressing past the metaphase stage (Dhyani et al., 2022), steroids weaken membrane integrity which leading to cell rupture (Dogan et al., 2017), tannins and saponins disrupt cell wall permeability which hindering bacterial growth (Nugrahani et al., 2025). Together, these compounds work synergistically to effectively combat pathogenic bacterial activity.

The decline in performance observed in T1 and T2, both in CD and the VH:CD ratio, may be due to feed sensitivity reactions. By T3, the chickens begin to adapt to the changes in feed formulations, with optimization occurring at T4. While no single study has specifically reported this mechanism in poultry, Coucke (2018) confirmed that feed sensitivity reactions are increasingly recognized as an important cause of intestinal tract atrophy. Unfortunately, as feed sensitivity was not measured in this study, the significance of this finding remains unclear.

Microbial population

One way to measure animal health is by investigating the presence of both beneficial and pathogenic bacteria. The gastrointestinal tract (GIT) of chickens begins to be colonized by bacterial species shortly after hatching (Ballou et al., 2016). The Lactobacillus genus account for nearly 70% of the microbiota (Shang et al., 2018), and is primarily found in the duodenum and jejunum. In contrast, Salmonella is a pathogenic bacterium that commonly causes illness in chicken (Stamilla et al., 2021). In the present study, we focused on investigating the presence of lactic acid bacteria (LAB) and Salmonella in jejunum to determine the chicken health.

The microbial analysis presented in Table 5 revealed that T3 exhibited a significant increase in LAB populations and a significant reduction in Salmonella sp. compared to other treatments (p<0.05). This suggests a synergistic effect of DOM bioactive substances in DOM, which not

 

Table 5: Effect of DOM supplementation on histomorphology of jejunal small intestine.

Items	Dietary treatments Jejunum microbial (cfu/g)					Statistics
	T0	T1	T2	T3	T4	SEM	P-value
Lactic acid bacteria (LAB)	17.47 × 106 a	29.64 × 105 a	27.1 × 105 a	41.47 × 106 b	12.42 × 106 a	41.83 × 105	0.001
Salmonella sp.	4.33 × 102 b	2.01 × 102 ab	1.66 × 102 a	1.00 × 102 a	4.03 × 102 b	4.04 × 101	0.026

 

Note: Means within a row without a common superscript differ significantly (P < 0.05). T0 (Control) = 0% (basal diet without adding DOM), T1 = Control + 0.5% DOM, T2 = Control + 1.0% DOM, T3 = Control + 1.5% DOM, and T4 = Control + 2.0% DOM.

 

only increase LAB populations but also reduce Salmonella levels in the gastrointestinal tract (GIT). At doses of 1.0% (T2) and 1.5% (T3), DOM may provide the most optimal effect in enhancing the gut microbiota composition. A lower dose of DOM, such as in T1 (0.5% DOM), may not be effective enough in suppressing Salmonella, while higher doses (T4, 2.0% DOM) may be excessive or fail to provide additional significant effects. These findings align with those of (Sirajuddin et al., 2025) who reported that both the lowest and highest doses of Dayak onion extract resulted in higher level of Salmonella. Similarly, Hidayah et al. (2021) highlighted the strong antibacterial properties of Dayak onion extract, particularly its ability in suppressing the Salmonella activity.

On the other hand, T0 and T4 exhibit similar levels of LAB and Salmonella sp. level in the GIT. The findings indicate that DOM supplementation effectively increased the LAB population in the jejunum of native chickens. This finding aligns with Zhang et al. (2015), who reported that phytobiotics, such as fermented ginkgo leaves rich in bioactive flavonoids, can increase the population of LAB in the ileum at a dose of 50 mg/kg of feed. This effect is likely due to the ability of DOM to modulate the intestinal microflora balance by increasing the population of LAB while suppressing pathogenic bacterial growth. The growth of LAB in the chicken GIT is influenced by favorable environmental conditions, including temperature, pH, and nutrient content (Widodo et al., 2015), suggesting DOM is able to provide ideal temperature, pH, sufficient nutrients for LAB growth. However, no single studies have reported the ability of Dayak onion to modulate temperature and pH in chicken GIT, thus we speculate that DOM is able to provide more digestible nutrients for LAB.

The main nutritional components in diet required by LAB include readily fermentable carbohydrates and protein (Wang et al., 2021). LAB use amylase to break down soluble carbohydrates and sequentially hydrolyze proteins. First, proteinase generates peptides, followed by peptidase activity that produces amino acids (Widodo et al., 2015). Additionally, LAB presence in the gut suppresses pathogenic bacteria by producing lactic acid and acetic acid, which lower pH and create inhospitable environment for pathogenic bacteria (Prastujati et al., 2023). This further explains why T0 and T4 exhibit similar concentration of both LAB and Salmonella, that is likely due to competition for nutritional resources in the GUT.

Furthermore, we also speculate that T3 provides the optimal concentration for promoting the growth of LAB. At this concentration, DOM might support LAB activity without overwhelming the gut environment with excess nutrients, creating a more favorable balance. In contrast, higher concentrations (T4 with 2.0% DOM) might not significantly improve LAB growth, or could even cause a shift in the microbial environment that negatively impacts LAB colonization, possibly due to changes in the gut’s overall nutrient availability or pH balance. However, Yuanita et al. (2019) emphasized that Dayak onion is able to lower pH in the GIT by producing short chain fatty acid. Thus, these findings remain unclear.

CONCLUSION

This study successfully evaluated the bioactive substance content of Dayak Onion Meal (DOM) and assessed its effect on the performance and intestinal health of superior native chicken. The results demonstrated that DOM supplementation, particularly T3 (1.5% DOM), positively influenced gut microbiota. Additionally, histomorphological changes were observed, suggesting potential benefits for intestinal health. While no significant improvements in overall growth performance were noted, these findings highlight the potential of DOM as a functional feed additive, offering a natural, antibiotic-free option for promoting gut health and modulating the gut microbiota in poultry.

Future studies should explore a wider range of DOM concentration and investigate its impact over longer feeding periods to fully understand its potential for improve native chicken health and performance. Additionally, research should examine the presence and impact of antinutritional factors (such as tannins and saponins) in DOM, particularly for their effect on nutrient absorption in native chicken.

ACKNOWLEDGEMENT

We would like to express our gratitude to the Faculty of Animal Science, Universitas Gadjah Mada, for funding support through the Graduate Research Grant Program number 1770/J01.1.25/PASCA.2020.

Novelty Statement

The novelty of this study lies in its first-time investigation of Dayak Onion Meal (DOM) supplementation in Superior Native Chicken (KUB) diets, focusing on comprehensive phytochemical profiling and its impacts on gut health parameters. Unlike previous research limited to broilers, quails, or meat preservation, this work uniquely demonstrates that 1.5% DOM optimally enhances lactic acid bacteria populations, suppresses Salmonella spp., and modulates jejunal histomorphology without compromising growth performance. This integrative approach provides new insights into DOM’s functional role as a natural, antibiotic-free feed additive tailored for indigenous poultry breeds, supporting sustainable poultry production in the post-antibiotic era.

AUTHOR’s CONTRIBUTION

Azhar Syafiq Imanullah played a central role in the study by designing and conducting the experiments, performing comprehensive laboratory analyses, interpreting the resulting data, and drafting the initial version of the manuscript. Nanung Danar Dono provided academic supervision throughout the research process, offering critical guidance during the experimental stages and making substantial revisions to enhance the quality of the manuscript. Bambang Ariyadi was actively involved in executing the experimental procedures and also made meaningful contributions to the development and refinement of the manuscript content. Aan Andri Yano contributed significantly to the structural and editorial development of the manuscript, ensuring coherence and clarity. All authors participated in the critical review of the manuscript and collectively approved the final version for submission.

Conflict of interest

We certify that there is no conflict of interest with any financial, personal, or other relationships with other people or organization related to the material discussed in the manuscript.

REFERENCES

Abdollahi MR, Zaefarian F, Ravindran V (2018). Feed intake response of broilers: Impact of feed processing. Anim. Feed Sci. Technol., 237: 154–165. https://doi.org/10.1016/j.anifeedsci.2018.01.013

Alghirani MM, Chung ELT, Jesse FFA, Sazili AQ, Loh TC (2021). Could phytobiotics replace antibiotics as feed additives to stimulate production performance and health status in poultry? An overview. J. Adv. Vet. Res., 11(4): 254–265.

Altaf M, Naveena BJ, Venkateshwar M, Kumar EV, Reddy G (2006). Single step fermentation of starch to L(+) lactic acid by Lactobacillus amylophilus GV6 in SSF using inexpensive nitrogen sources to replace peptone and yeast extract- Optimization by RSM. Process Biochem., 41(2): 465–472. https://doi.org/10.1016/j.procbio.2005.07.011

Babaei Rad S, Mumivand H, Mollaei S, Khadivi A (2025). Effect of drying methods on phenolic compounds and antioxidant activity of Capparis spinosa L. fruits. BMC Plant Biol., 25(1): 1–14. https://doi.org/10.1186/s12870-025-06110-y

Ballou AL, Ali RA, Mendoza MA, Ellis JC, Hassan HM, Croom WJ, Koci MD (2016). Development of the chick microbiome: How early exposure influences future microbial diversity. Front. Vet. Sci., 3(2): 1–12. https://doi.org/10.3389/fvets.2016.00002

Berliana, Nurhayati, Nelwida (2018). Substitution garlic with black garlic in the ration on early egg production of layer quail (Coturnix - coturnix japonica). J. Agripet, 18(2): 95–102. https://doi.org/10.17969/agripet.v18i2.12779.

Çalişlar S (2018). Effects of tannins on poultry nutrition. J. Agric. Nat., 21(4): 615–623.

Classen HL (2017). Diet energy and feed intake in chickens. Anim. Feed Sci. Technol., 233: 13–21. https://doi.org/10.1016/j.anifeedsci.2016.03.004

Coucke F (2018). Food intolerance in patients with manifest autoimmunity. Observational study. Autoimmun. Rev., 17(11): 1078–1080. https://doi.org/10.1016/j.autrev.2018.05.011

Dhyani P, Quispe C, Sharma E, Bahukhandi A, Sati P, Attri DC, Szopa A, Sharifi-Rad J, Docea AO, Mardare I, Calina D, Cho WC (2022). Anticancer potential of alkaloids: a key emphasis to colchicine, vinblastine, vincristine, vindesine, vinorelbine and vincamine. Cancer Cell Int., 22(1): 1–20. https://doi.org/10.1186/s12935-022-02624-9

Dogan A, Otlu S, Çelebi Ö, Kiliçle PA, Saglam AG, Dogan ANC, Mutlu N (2017). An investigation of antibacterial effects of steroids. Turk. J. Vet. Anim. Sci., 41(2): 302–305. https://doi.org/10.3906/vet-1510-24

Dono ND (2012). Nutritional Strategies To Improve Enteric Health and Growth Performance of Poultry in the Post Antibiotic Era (Doctoral thesis). Glasgow, Scotland : University of Glasgow. Retrived June 08, 2025, from https://theses.gla.ac.uk/4025/1/2012DonoPhD.pdf

Fitriansyah SN, Puteri N, Muthmainnah N, Setiani NA (2021). Kandungan kimia dan aktivitas antioksidan pada umbi bawang dayak (Eleutherine palmifolia (L.) Merr.) hasil fermentasi. J. Sains Teknol. Farm. Indonesia, 10(2): 44. https://doi.org/10.58327/jstfi.v10i2.181

Gautam H, Ayalew LE, Shaik NA, Subhasinghe I, Popowich S, Chow-Lockerbie B, Dixon A, Ahmed KA, Tikoo SK, Gomis S (2024). Exploring the predictive power of jejunal microbiome composition in clinical and subclinical necrotic enteritis caused by Clostridium perfringens: insights from a broiler chicken model. J. Transl. Med., 22(1): 1–18. https://doi.org/10.1186/s12967-023-04728-w

Gheisar MM, Zhao P, Kim IH (2018). Addition of phytogenic blend in different nutrient density diets of meat-type ducks. J. Appl. Anim. Res., 46(1): 854–859. https://doi.org/10.1080/09712119.2017.1411265

Haerul M, Hermana W, Sumiati (2024). The effect of feeding of eleutherine palmifolia meal in diet on the hematological profile of quail. Anim. Nutr. Feed Technol., 22(2): 72–78. https://doi.org/10.29244/jintp.22.2.72-78

Haque M, McKimm J, Sartelli M, Dhingra S, Labricciosa FM, Islam S, Jahan D, Nusrat T, Chowdhury TS, Coccolini F, Iskandar K, Catena F, Charan J (2020). Strategies to prevent healthcare-associated infections: A narrative overview. Risk Manage. Healthc. Policy, 13: 1765–1780. https://doi.org/10.2147/RMHP.S269315

Hasnat, H, Shompa SA, Islam MM, Alam S, Richi FT, Emon NU, Ashrafi S, Ahmed NU, Chowdhury MNR, Fatema N, Hossain MS, Ghosh A, Ahmed F (2024). Flavonoids: A treasure house of prospective pharmacological potentials. Heliyon, 10(6): e27533. https://doi.org/10.1016/j.heliyon.2024.e27533

Hibbard R, Chapot L, Yusuf H, Ariyanto KB, Maulana KY, Febriyani W, Cameron A, Vergne T, Faverjon C, Paul MC (2023). It’s a habit. They’ve been doing it for decades and they feel good and safe. A qualitative study of barriers and opportunities to changing antimicrobial use in the Indonesian poultry sector. PLoS One, 18(9 September): 1–22. https://doi.org/10.1371/journal.pone.0291556

Hidayah R, Purwanti S, Jamilah J (2021). Anti-bacterial activity of Dayak onions extract (Eleutherine palmifolia) against Salmonella spp and Escherichia coli. IOP Conf. Ser. Earth Environ. Sci., 788(1): 1–6. https://doi.org/10.1088/1755-1315/788/1/012069

Hidayat C, Asmarasari SA (2015). Native chicken production in Indonesia: A review. Indones. J. Anim. Sci., 17(1): 1-11. https://doi.org/10.25077/jpi.17.1.1-11.2015

Jayanegara A, Yogianto, Wina E, Sudarman A, Kondo M, Obitsu T, Kreuzer M (2020). Combination effects of plant extracts rich in tannins and saponins as feed additives for mitigating. Animals, 10: 1–14. https://doi.org/10.3390/ani10091531

Kavoi BM, Gakuya DW, Mbugua PN, Kiama SG (2016). Effects of dietary Moringa oleifera leaf meal supplementation on chicken intestinal structure and growth performance. J. Morphol. Sci., 33(4): 186–192. https://doi.org/10.4322/jms.095915

Kiela PR, Ghishan FK (2016). The physiology of intestinal absorption. Best Prac. Res. Clin. Gastroenterol., 30(2): 145–159. https://doi.org/10.1016/j.bpg.2016.02.007

Lisnahan CV, Nahak OR (2020). Growth performance and small intestinal morphology of native chickens after feed supplementation with tryptophan and threonine during the starter phase. Vet. World, 13(12): 2765–2771. https://doi.org/10.14202/vetworld.2020.2765-2771

Marchewka J, Sztandarski P, Zdanowska-Sąsiadek Ż, Adamek-Urbańska D, Damaziak K, Wojciechowski F, Riber AB, Gunnarsson S (2021). Gastrointestinal tract morphometrics and content of commercial and indigenous chicken breeds with differing ranging profiles. Animals, 11(7): 1–15. https://doi.org/10.3390/ani11071881

Mehdi Y, Létourneau-Montminy MP, Gaucher ML, Chorfi Y, Suresh G, Rouissi T, Brar SK, Côté C, Ramirez AA, Godbout S (2018). Use of antibiotics in broiler production: Global impacts and alternatives. Anim. Nutr., 4(2): 170–178. https://doi.org/10.1016/j.aninu.2018.03.002

Miles RD, Butcher GD, Henry PR, Littell RC (2006). Effect of antibiotic growth promoters on broiler performance, intestinal growth parameters, and quantitative morphology. Poult. Sci., 85(3): 476–485. https://doi.org/10.1093/ps/85.3.476

Monasdir, Nugroho AA, Riskayanti, Husein F, Nisfimawardah L, Astuti E, Setyowati EAH, Nio MM (2024). Physical and chemical properties of chicken sausage with the addition of dayak onion (Eleutherine palmifolia (L) Merr) from Central Kalimantan. J. Ilmu Teknol. Hasil Ternak, 19(2): 69–77. https://doi.org/10.21776/ub.jitek.2024.019.02.1

Mukti YP, Yusup B, Dewi ADR, Kang SC (2023). Fortification of dayak onion extract (Eleutherine palmifolia (L.) Merr.) to jelly drink as a functional food. Adv. Biol. Sci. Res., 2022: 64–72. https://doi.org/10.2991/978-94-6463-322-1_8

Muslim C, Agustian A, Perdana RP, Aldillah R, Darwis V (2021). Constraints and benefits of KUB chicken cultivate in Central Java Province, Indonesia. E3S Web Conf., 306: 1–8. https://doi.org/10.1051/e3sconf/202130602027

Nugrahani NA, Nurilyana MM, Faizal IA, Kholifa M, Hafizi I (2025). Efficacy of avocado seed extract in preventing, inhibiting, and eliminating Prevotella intermedia biofilms: An in vitro study. Vet. World, 18(2): 408–418. https://doi.org/10.14202/vetworld.2025.408-418

Obianwuna UE, Chang X, Oleforuh-Okoleh VU, Onu PN, Zhang H, Qiu K, Wu S (2024). Phytobiotics in poultry: revolutionizing broiler chicken nutrition with plant-derived gut health enhancers. J. Anim. Sci. Biotechnol., 15(1): 1–33. https://doi.org/10.1016/j.psj.2024.103952

Oladeji OM, Mugivhisa LL, Olowoyo JO (2025). Antibiotic residues in animal products from some African countries and their possible impact on human health. Antibiotics, 14(90): 1–5. https://doi.org/10.3390/antibiotics14010090

Pereira DM, Valentão P, Pereira JA, Andrade PB (2009). Phenolics: From chemistry to biology. Molecules, 14(6): 2202–2211. https://doi.org/10.3390/molecules14062202

Prastujati AU, Amaniyah M, Priyadi DA, Arief II, Fernadi G (2023). Identification of pathogenic bacteria in fermented bekamal using B14A isolate. IOP Conf. Ser.: Earth Environ. Sci., 1168(1): 1–6. https://doi.org/10.1088/1755-1315/1168/1/012043

Rahayu HSP, Takdis M, Asnidar, Dewi M, Rameda NM, Haryono P (2021). Response and feasibility of KUB chicken on household business level in rural area Sigi district. Adv. Econ. Bus. Manage. Res., 199: 46–52.

Ravindran V, Reza AM (2021). Nutrition and digestive physiology of the broiler chick: State of the art and outlook. Animals, 11: 1–23. https://doi.org/10.3390/ani11102795

Riadh DN, Paramita S, Yadi (2024). Application of dayak onion (Eleutherine bulbosa Urb.) as a preservative for animal protein: chicken meat. Int. J. Multidis. Appl. Bus. Educ. Res., 5(2): 375–382. https://doi.org/10.11594/ijmaber.05.02.01

Romadhon T, Rahayu WP, Kusumanigrum HD (2023). Ekstrak bawang dayak penghambat pertumbuhan Escherichia coli dan Staphylococcus aureus pada daging ayam. J. Ilmu Pertan. Indonesia, 28(3): 504–512. https://doi.org/10.18343/jipi.28.3.504

Ronquillo MG, Hernandez JCA (2017). Antibiotic and synthetic growth promoters in animal diets: Review of impact and analytical methods. Food Contr., 72: 255–267. https://doi.org/10.1016/j.foodcont.2016.03.001

Salehi TZ, Mahzounieh M, Saeedzadeh A (2005). The isolation of antibiotic-resistant Salmonella from intestine and liver of poultry in Shiraz Province of Iran. Int. J. Poult. Sci., 4(5): 320–322. https://doi.org/10.3923/ijps.2005.320.322

Sekarsari RA, Damanhuri D, Kuswanto K (2024). Exploration of the dayak onion (Eleutherine palmifolia (L.) Merr) in the Borderlands of North Kalimantan, Indonesia. Agro Bali Agric. J., 7(2): 457–466. https://doi.org/10.37637/ab.v7i2.1659

Shang Y, Kumar S, Oakley B, Kim WK (2018). Chicken gut microbiota: Importance and detection technology. Front. Vet. Sci., 5: 1–13. https://doi.org/10.3389/fvets.2018.00254

Silitonga L, Wibowo S, Bangun EB (2020). The effect of giving dayak union flour (Eleutherine palmifolia Merr.) on organs morphometric and meat fat of broiler chicken. Ziraa’ah, 45(1): 10–20. https://doi.org/10.31602/zmip.v45i1.2263

Sirajuddin MM, Rusman R, Widodo W, Suryanto E (2025). Evaluation of the antibacterial activity of dayak onion extract (Eleutherine palmifolia L.) against pathogenic Salmonella spp. and Escherichia coli. Adv. Anim. Vet. Sci., 13(7): 1517–1524. https://doi.org/10.17582/journal.aavs/2025/13.7.1517.1524

Stamilla A, Ruiz-Ruiz S, Artacho A, Pons J, Messina A, Randazzo CL, Caggia C, Lanza M, Moya A (2021). Analysis of the microbial intestinal tract in broiler chickens during the rearing period. Biology, 10(9): 1–19. https://doi.org/10.3390/biology10090942

Supomo, Syamsul ES, Apriliana A, Saleh C, Erwin, Lestari D (2019). Antioxidant assay of dayak onion (Eleutherine palmifolia) via dpph (1,1-difenil-2-pikrilhidrazil) and BSLT test for its active fraction. Rasayan J. Chem., 12(3): 1340–1346. https://doi.org/10.31788/RJC.2019.1235264

Susilawati NM, Bria M, Foekh NP (2022). Inhibitory test of Dayak onion extract (Eleutherine palmifolia (L) merragainst gram negative and gram positive bacteria. Sci. Midwifery, 10(4): 2972–2976. https://doi.org/10.35335/midwifery.v10i4.745

Tsado D, Ayanwale BA, Ijaiya AT, Egwim EC (2019). Effects of cellulase and pectinase hydrolyzed corncob based diets on performance and carcass yield of broiler chickens. Niger. J. Anim. Sci., 21(1): 44–55.

Valli C, Kavitha R (2023). Phytobiotics as an alternative feed additive. Indian J. Vet. Anim. Sci. Res., 52(4): 1–14.

Wanderley R de OS, Figueirêdo RMF de, Queiroz AJ de M, Santos FS dos, Silva AP de F, Paiva YF, Moura HV, Silva ET de V, Carvalho AJ de BA, Lima M dos S, Campos ARN, Gregório MG, Lima AGB de (2023). Effect of drying temperature on antioxidant activity, phenolic compound profile and hygroscopic behavior of pomegranate peel and seed flours. LWT Food Sci. Technol., 189: 115514. https://doi.org/10.1016/j.lwt.2023.115514

Wang Y, Wu J, Lv M, Shao Z, Hungwe M, Wang J, Bai X, Xie J, Wang Y, Geng W (2021). Metabolism characteristics of lactic acid bacteria and the expanding applications in food industry. Front. Bioeng. Biotechnol., 9: 1–19. https://doi.org/10.3389/fbioe.2021.612285

Widodo TS, Sulistiyanto B, Utama CS (2015). The amount of lactic acid bacteria in ileal digesta and secal digesta of broiler fed ration containing fermented abandoned feed. J. Agripet, 15(2): 98–103. https://doi.org/10.17969/agripet.v15i2.2376

Xu C, Kong L, Gao H, Cheng X, Wang X (2022). A review of current bacterial resistance to antibiotics in food animals. Front. Microbiol., 13: 1–16. https://doi.org/10.3389/fmicb.2022.822689

Yano AA, Suprayogi WPS, Darmawan MA, Saputro WS, Saraswati PN, Setyono W, Akhirini N (2024). Effect of thermo-mechanically fermented soybean meal on nutrient digestibility and carboxymethyl cellulase activity of jawa super chicken. Indones. J. Anim. Sci., 34(3): 286–298. https://doi.org/10.21776/ub.jiip.2024.034.03.1

Yuanita I, Sunarti D, Wahyuni HI, Suthama N (2019). Feeding Dayak onion (Eleutherine palmifolia) extract and Lactobacillus acidophilus mixture on blood biochemicals, meat quality characteristics and growth performance in broiler chickens. Livest. Res. Rural Dev., 31(9): 1–11.

Yuswi NCR (2017). Antioxidant Extraction of Bawang Dayak (Eleutherine palmifolia) with ultrasonic bath (study type of solvent and extraction time). J. Pangan Agroind., 5(1): 71–79.

Zakiyatulyaqin Z, Setiawan D, Purnomosidi M (2020). The performance of broiler chickens with the addition of dayak onion extract in drinking water. J. Ternak, 11(2): 91–96. https://doi.org/10.30736/jt.v11i2.77

Zhang XH, Sun ZY, Cao FL, Ahmad H, Yang XH, Zhao LG, Wang T (2015). Effects of dietary supplementation with fermented ginkgo leaves on antioxidant capacity, intestinal morphology and microbial ecology in broiler chicks. Br. Poult. Sci., 56(3): 370–380. https://doi.org/10.1080/00071668.2015.1030590