Open Veterinary Journal, (2025), Vol. 15(9): 4681-4690

Research Article

10.5455/OVJ.2025.v15.i9.72

Water hyacinth: A promising functional feed ingredient to optimize reproductive performance in Garut rams

Aeni Nurlatifah¹, Kurniawan Dwi Prihantoko^1,2, Jean Baptiste Menassol³, Herdis Herdis², Athhar Manabi Diansyah^4*, Faheem Ahmed Khan⁵ and Mashitah Shikh Maidin⁶

¹Faculty of Animal Science, Gadjah Mada University, Depok, Indonesia

²Research Center for Animal Husbandry, National Research and Innovation Agency, Bogor, Indonesia

³SELMET, Institut Agro, INRAE, CIRAD, Univ Montpellier, Montpellier, France

⁴Faculty of Animal Science, Hasanuddin University, Makassar, Indonesia

⁵Department of Transfusion Medicine and Clinical Microbiology, Faculty of Allied Health Sciences, Chulalongkorn University, Bangkok, Thailand

⁶Department of Biology Faculty of Science, Universiti Putra Malaysia, Seri Kembangan, Malaysia

*Corresponding Author: Athhar Manabi Diansyah. Faculty of Animal Science, Hasanuddin University, Makassar, Indonesia. Email: athhar.md [at] unhas.ac.id

Submitted: 14/04/2025 Revised: 27/07/2025 Accepted: 14/08/2025 Published: 30/09/2025

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Abstract

Background: Reproductive efficiency in Garut rams is crucial for sustainable sheep farming in West Java, but suboptimal performance has limited its application. This study explores the use of WH as a functional feed additive to enhance semen quality and hormonal regulation, offering an innovative approach to optimize reproduction and address environmental concerns.

Aim: This study examined the impact of a WH (Eichhornia crassipes)-based diet on fertility in Garut rams, focusing on semen quality, sperm kinematics, hormonal responses, and reproductive capacity.

Methods: This study involved ten Garut rams fed a formulated diet for 90 days, with semen parameters (motility, viability, abnormalities, acrosome integrity, and kinematics) assessed every 15 days using computer-assisted sperm analysis. Hormonal levels [testosterone, follicle-stimulating hormone (FSH), and luteinizing hormone (LH)] were measured via enzyme-linked immunosorbent assay, and conception rates were evaluated after artificial insemination.

Results: Statistical analysis showed significant improvements in semen quality, motility, progressive motility, and acrosome integrity from day 60 to 90. Sperm viability also showed a progressive increase, with significant differences observed from day 0 to 30, continuing to rise through day 90. Sperm abnormalities gradually increased, but acrosome integrity demonstrated the most significant enhancement, reaching its peak at day 90. Sperm kinematic parameters, such as curve linear velocity, average path velocity, and straight line velocity, were significantly higher on day 90 than on day 0. Hormonal analysis revealed a marked increase in testosterone levels from day 30, while FSH and LH levels remained stable throughout the study. Conception rates increased progressively, with the highest values recorded at days 60 and 90.

Conclusion: A WH-based diet can enhance the reproductive performance of Garut rams, offering a sustainable feeding strategy that boosts breeding efficiency while helping to manage this invasive plant. Future studies covering longer time frames, larger and more diverse flocks, and on-farm conditions will be essential to confirm these benefits and optimize ration formulations.

Keywords: Garut rams, Water hyacinth, Semen quality, Sperm kinematics, Hormonal regulation.

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Introduction

The Garut rams (Ovis aries) hold significant cultural, economic, and ecological value in Indonesia, particularly in West Java. These rams are highly regarded not only for their resilience and adaptability to diverse environmental conditions but also for their exceptional reproductive performance (Nurlatifah et al., 2023). Garut rams are an integral part of local agricultural practices and cultural traditions, and events such as the Garut sheep festival celebrate their importance. In addition to their role in meat production, Garut rams are a key asset in breeding programs aimed at improving livestock quality and maintaining the region’s rich sheep farming heritage. The continued preservation and enhancement of the genetic traits of Garut rams are crucial for sustaining both the local economy and cultural practices tied to these animals (Herdis et al., 2019).

Despite their importance, the reproductive performance of Garut rams in breeding programs has yet to reach its full potential. Insufficient knowledge about optimal feeding strategies and the impact of nutritional interventions on semen quality and fertility hinders the effective management of breeding programs. Although Garut rams are bred for their superior traits, reproductive challenges persist, including variations in sperm quality and hormonal imbalances. Previous studies have reported suboptimal semen quality in some populations, with average sperm motility below 60%, viability under 70%, and increased abnormalities exceeding 15%, which can compromise fertility outcomes (Ariyanto et al., 2020). These issues emphasize the need for innovative approaches to improve breeding efficiency and optimize the genetic potential of Garut rams, thereby enhancing the local sheep farming industry’s productivity and sustainability.

The use of functional feed ingredients is a promising strategy for improving reproductive performance. Water hyacinth (WH) (Eichhornia crassipes) has the potential to be a sustainable feed ingredient due to its high protein content, favorable nutrient profile, and year-round availability (Song et al., 2025). Although considered an invasive weed in Indonesia, its use in livestock diets remains underutilized (Nandiyanto et al., 2023). Studies in cattle have shown that plant-based feeds, such as WH, Azolla pinnata, and Leucaena leucocephala, can improve reproductive performance, including semen quality and hormonal profiles (Isnawati et al., 2023; Ojediran et al., 2024). However, research on small ruminants, especially Garut rams, is limited. Further studies are needed to determine optimal inclusion levels, address anti-nutritional factors, and evaluate long-term reproductive effects to support both sustainable feeding strategies and invasive species control. This study aimed to evaluate the impact of WH-based diets on the reproductive performance of Garut rams, particularly in terms of semen quality and hormonal responses, aligning with previous research on other species while focusing on species-specific parameters relevant to rams.

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Materials and Methods

Experimental design

In this experiment, 10 Garut rams were selected based on similar characteristics, including age (~23 months) and body weight (~40 kg), to ensure uniformity in physiological and reproductive maturity. Rams aged approximately 23 months are considered sexually mature with stable hormonal profiles and fully developed reproductive organs, making them suitable for evaluating semen quality and fertility-related parameters. A body weight of approximately 40 kg reflects a healthy, mature ram in good condition, minimizing variability due to nutritional or developmental differences that could influence the study outcomes. The rams were fed WH (E. crassipes) for 90 days. Each individual animal was kept in approximately 1.5 m², equipped with an environmentally controlled slatted floor facility. Throughout the experimental period, the animals had free access to feed and water, which were provided via a self-feeder and nipple drinker, respectively. The temperature and humidity within the animal housing were controlled within appropriate ranges: the temperature was maintained between 16°C and 25°C, and the humidity was maintained at 65%. The animals were maintained according to the guidelines for the husbandry of male sheep. The reproductive performance of the rams was evaluated during the feeding period, with observations made every 15 days.

Feed formulation and nutritional quality

In this study, feed formulations were prepared for rams over a 90-day feeding period. The feed formulas contain WH, soybean meal, corn flour, rice bran, coconut meal, fish meal, and premix. Each ingredient is given a specific proportion according to the formulation (Table 1).

Dried WH was included at 30% because of its high protein content, wide availability, and safe inclusion level regarding potential anti-nutritional factors. Rice bran (25%) and coconut meal (20%) were used as energy and fiber sources to support rumen fermentation. Soybean meal (6%) and fish meal (4%) provided high-quality protein and essential amino acids critical for spermatogenesis. Corn flour (14.5%) contributed readily digestible carbohydrates to meet metabolic energy demands, while a premix (0.5%) was added to ensure adequate intake of essential vitamins and minerals necessary for optimal reproductive function. This formulation was developed to support reproductive performance and evaluate the potential of WH as a sustainable feed ingredient.

The preparation process involves weighing the ingredients, thoroughly mixing them, and storing the prepared feed until it is ready to be fed to the rams. The feed was subjected to nutritional quality testing five times following the procedures outlined (Irawan et al., 2022; Azis et al., 2023) using proximate analysis (Table 2).

Table 1. Composition of feed ingredients in dried WH and dried azolla formulations.

Table 2. Nutritional quality of dried WH and dried azolla formulations.

The feed’s nutritional composition meets the requirements specified in SNI 8819-2019, which sets the standards for sheep feed for breeding.

Evaluation and processing of sperm

Semen collection from Garut rams was conducted weekly using an artificial vagina. The quality of sperm was assessed based on motility, progressive motility, kinematics, abnormality, viability, and acrosome integrity (Diansyah et al., 2023). Motility, progressive motility, and kinematics were analyzed by placing a sperm sample on a glass object and using a computer-assisted sperm analysis (CASA) system (Vision Version 3.7.5, Minitub, Germany) (Diansyah et al., 2021). The measured kinematic parameters included curve linear velocity (VCL), average path velocity (VAP), and straight line velocity (VSL) (Rahmat et al., 2024). Viability and abnormality were evaluated through 2% eosin staining under a trinocular microscope at 400× magnification, where dead sperm appeared red, and abnormalities such as severed tails, broken tails, and irregular head shapes were recorded (Surahman et al., 2021). Acrosome integrity was examined using a formol-saline solution, with intact acrosomes appearing as black head tips under a trinocular microscope (Diansyah et al., 2022a). Semen was frozen using a commercial extender (Andromed, Germany), equilibrated at 5°C for 4 hour, and then cryopreserved in liquid nitrogen vapor for 15 minute before storage at −196°C (Diansyah et al., 2022b).

Enzyme-linked immunosorbent assay

The enzyme-linked immunosorbent assay (ELISA) was used to determine the concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and testosterone in Garut rams. Blood samples were collected via jugular venipuncture in the morning, coinciding with semen collection. The samples were then centrifuged at 3,000 rpm for 15 minute to separate the serum, which was subsequently stored at −20°C until analysis (Ma et al., 2018). The storage temperature of 20°C was chosen to preserve hormone stability and prevent degradation or denaturation of serum proteins prior to analysis. A commercially available ELISA kit specific to each hormone was used following the manufacturer’s instructions for rams. To initiate a colorimetric reaction, serum samples and standards were added to 96-well microplates pre-coated with hormone-specific antibodies, incubated, and treated with an enzyme-labeled secondary antibody and substrate solution. The reaction was stopped using sulfuric acid, and the absorbance was measured at 450 nm using a microplate reader. Hormone concentrations were calculated based on standard curves generated for each hormone, with all samples analyzed in duplicates to ensure accuracy, and results were expressed in ng/mL (Yin et al., 2018; Oematan et al., 2023).

Reproductive capacity assessment

The reproductive capacity of rams was evaluated based on their conception rate using artificial insemination (AI ) with semen from rams that had been given specific feed treatments. Semen frozen from each treatment per observation per ram was then inseminated into 12 ewes. The selection of 12 ewes per treatment group was based on standard reproductive trial designs aimed at providing sufficient statistical power to detect treatment effects while considering logistical and ethical constraints related to animal use. This sample size balances the need for reliable reproductive performance data with the practical limitations of managing synchronized estrus and AI procedures in small ruminants. To determine conception rates, pregnancy diagnosis was performed at least 45 days post-insemination using transabdominal ultrasonography or rectal palpation (Yusuf et al., 2024).

Statistical analysis

The observed parameters included sperm quality, sperm kinematics, hormone levels, and reproductive performance. Data were analyzed using parametric statistical methods. All parameter values were tested for normality and homogeneity of variances prior to statistical analysis. Because the data were normally distributed, comparisons between feed treatment groups for sperm quality, hormone levels, and reproductive performance were performed using the paired sample t-test. To enhance the accuracy and reliability of the statistical results, data were expressed as mean ± SEM, and 95% confidence intervals (CIs) were calculated to indicate the precision of estimates. Differences were considered statistically significant at p < 0.05. All statistical analyses were performed using SPSS version 25 for Windows (IBM Corp., Chicago, IL).

Ethical approval

The Animal Ethics Committee of the National Research and Innovation Agency, Indonesia, approved the experimental protocol (approval no. 204/KE.02/SK/11/2023).

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Results

Sperm quality and kinematics in Garut rams during hyacinth water-feeding periods

Figure 1 illustrates the changes in sperm quality parameters regarding motility, progressive motility, viability, abnormality, and acrosome integrity over a 90-day period in Garut rams during WH feeding periods. Significant differences (p < 0.05) between days are indicated. Motility showed no significant differences between days 0 and 15. Values at days 60 (76.57%), 75 (78.91%), and 90 (79.67%) were significantly higher compared to earlier days. Significant improvements were observed from day 60 (76.57%) and continued through to day 90 (79.67%), indicating a gradual increase in motility after day 60. Significant differences were observed between day 0 (50.51%) and day 15 (49.48%) for progressive motility. From day 30, values showed continuous improvement, from day 30 (53.33%) and continued increasing to day 45 (56.03%), day 60 (57.26%), day 75 (58.45%), and day 90 (59.85%). The highest values were observed on day 90, which were significantly greater than those on earlier days, suggesting progressive enhancement over the study period. Viability showed a significant increase from day 0 (81.28%) to day 30 (83.56%), with further improvements between days 45 (84.84%) and 60 (85.97%). Values at days 75 (87.65%) and 90 (88.73%) were significantly higher than those at earlier days, reflecting consistent improvement in sperm viability throughout the observation period. The abnormality exhibited a significant increase from day 0 (6.48%) to day 15 (6.21%), and further significant differences were observed from day 30 (6.50%) onward. Values at days 60 (7.32%), 75 (7.69%), and 90 (7.66%) were significantly higher than those at earlier stages, indicating a gradual increase in sperm abnormalities over the study duration. This rising trend suggests a potential side effect of prolonged WH feeding on sperm morphology.

Acrosome integrity demonstrated significant improvements starting from day 15 (80.77%), with significantly higher values at days 30 (81.97%), 45 (82.51%), 60 (83.97%), 75 (85.22%), and 90 (85.88%) than at day 0 (80.15%). The highest values were recorded at day 90, showing a marked improvement in acrosome integrity, reflecting enhanced sperm head protection over time. Figure 1 presents significant changes in all parameters, with consistent improvements in acrosome integrity and viability over time. Motility and progressive motility demonstrated initial increases followed by stabilization, whereas the abnormality gradually increased throughout the observation period.

Figure 2 illustrates the sperm kinematic parameters regarding VCL, VAP, and VSL in Garut rams during the WH feeding periods over a 90-day observation period. Significant differences (p < 0.05) between days are indicated. Significant increases in VCL were observed starting from day 15 (95.17 µm/s) onward. By day 30 (100.13 µm/s), the values were significantly higher than those at day 0 (90.20 µm/s), and this trend continued with further significant increases at days 45 (105.05 µm/s), 60 (104.98 µm/s), 75 (115.22 µm/s), and 90 (119.90 µm/s), where the values reached their peak, reflecting enhanced sperm motility.

Fig. 1. Sperm quality in Garut rams during WH feeding periods; different superscripts in the same line indicate significant differences at p < 0.05.

Fig. 2. Sperm kinematics in Garut rams during WH feeding periods; different superscripts in the same line indicate significant differences at p <0.05.

The VAP showed similar trends, with significant increases starting from day 15 (61.68 µm/s), and values at days 30 (63.07 µm/s), 45 (64.71 µm/s), 60 (66.41 µm/s), and 75 (68.45 µm/s) were significantly higher than at day 0 (60.23 µm/s). By day 90 (69.76 µm/s), values were significantly higher than those in earlier days, indicating a sustained improvement in VAP over the study period. The VSL exhibited a continuous rise from day 0 (30.18 µm/s) to day 90 (39.77 µm/s), with significant differences observed at each time point. The intermediate values were recorded on days 15 (31.84 µm/s), 30 (33.07 µm/s), 45 (34.90 µm/s), 60 (36.59 µm/s), and 75 (38.33 µm/s). The lowest values were recorded on day 0, whereas the highest values were recorded on day 90, suggesting consistent improvement in straight-line velocity throughout the observation period. Figure 2 demonstrates a consistent increase in all sperm kinematic parameters, with significant improvements in VCL, VAP, and VSL over the 90-day period of WH feeding in Garut rams.

Hormone level in Garut rams during hyacinth-feeding periods

Figure 3 illustrates the hormone levels—FSH, LH, and testosterone measured in Garut rams during the WH feeding periods over a 90-day observation period. Significant differences (p < 0.05) between days are indicated. A significant increase in testosterone levels was observed starting from day 30, with the highest values recorded at day 90. Values at days 30, 60, 75, and 90 were significantly higher than those at day 0, suggesting a sustained elevation in testosterone levels throughout the study period. In contrast, FHS and LH levels remained relatively stable throughout the 90-day study period, with no significant differences observed between the days.

Reproductive capacity in Garut rams during hyacinth-feeding periods

Figure 4 presents the conception rate in Garut rams during the WH feeding periods over a 90-day observation period. Significant differences (p < 0.05) between days are indicated. Initially, the conception rate was comparatively lower at day 0, but it exhibited a progressive increase as the feeding period advanced.

Notably, by day 45, a significant improvement in the conception rate was observed by day 45 relative to days 0 and 15. The rate continued to increase through days 60, 75, and 90, with the highest values recorded at days 60 and 90.

Fig. 3. Hormonal levels in Garut rams during WH feeding periods; different superscripts in the same line indicate significant differences at p < 0.05.

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Discussion

The reproductive performance of Garut rams is crucial for sheep farming in West Java, Indonesia. However, challenges, such as suboptimal semen quality and hormonal imbalances, hinder the effectiveness of breeding programs. This study evaluated the effects of WH-based diets on semen quality, sperm kinematics, hormonal responses, and reproductive capacity, providing insights into how this alternative feed source can improve reproductive health in Garut rams.

WH supplies a spectrum of bioactive metabolites—including phenolics, flavonoids, and alkaloids—that confer antioxidant, anti-inflammatory, and endocrine-modulating effects pertinent to male reproduction. These compounds safeguard sperm DNA integrity, maintain membrane fluidity, and facilitate acrosome formation by reducing oxidative stress in testicular tissue. In particular, flavonoids modulate the hypothalamic–pituitary–gonadal axis and upregulate key steroidogenic genes such as Steroidogenic Acute Regulatory (StAR) protein and cytochrome P450 C17 (CYP17A1), thereby enhancing testosterone biosynthesis in Leydig cells. Complementary protein-rich ingredients (e.g., soybean and fish meal) supply amino-acid precursors—especially arginine and lysine—that stimulate nitric oxide production and endocrine signaling, further optimizing Leydig cell function. In addition, unsaturated fatty acids in the ration may promote steroidogenesis by elevating the expression or activity of enzymes such as 3β-hydroxysteroid dehydrogenase alongside StAR, collectively increasing testosterone output and improving spermatogenic performance.

The significant improvements in sperm quality parameters observed in this study, particularly motility, progressive motility, viability, acrosome integrity, and sperm abnormalities, can be attributed to the nutritional composition of the water hyacinth based (WHB) diet. The feed formulation contained key ingredients, such as soybean meal, corn flour, and fish meal, which are rich in essential nutrients, such as proteins, fatty acids, and micronutrients crucial for sperm function (Table 2) (Kujoana et al., 2024). The nutritional quality of the feed, particularly the crude protein and crude fat contents, may support cellular energy production, membrane stability, and overall sperm health (Srivastava et al., 2023). The addition of specific plant-based ingredients with balanced protein and fat contents can enhance sperm motility and viability (Cojkic et al., 2023). These findings agree with research on other livestock species, where plant-based feed inclusion has been linked to improved semen quality. The observed improvement in sperm motility and acrosome integrity after day 60 and 90, respectively, can be linked to the gradual accumulation of these nutrients in the system, allowing for the stabilization of sperm membranes and enhanced functional activity (Chao et al., 2023).

Fig. 4. Reproductive capacity in Garut rams during WH feeding periods; different superscripts indicate significant differences at p < 0.05.

The sperm quality parameters improved consistently throughout the feeding period with the WHB diet. Total and progressive motility increased following an initial adaptation phase, suggesting that WH—rich in antioxidants and essential fatty acids—emerges with sustained intake and supports mitochondrial function and membrane stability. Sperm viability and acrosome integrity also showed steady improvement, indicating enhanced membrane protection and fertilizing potential, likely due to phytochemicals in the diet’s cytoprotective properties. Although sperm abnormalities increased slightly over time, they remained within physiological limits and did not negate the overall improvements observed in other parameters, suggesting that the effect may stem from natural variation or handling stress rather than toxicity (Ratnasari et al., 2018). Collectively, these trends underscore the reproductive benefits of WH and demonstrate that its positive impact on sperm function becomes more apparent with prolonged supplementation, in line with previous findings on phytogenic feed ingredients. CASA was crucial in providing an objective and precise measurement of sperm kinematics, ensuring that sperm motility and velocity were optimally evaluated (Maulana et al., 2022). The kinematic parameters of VCL, VAP, and VSL showed significant improvements, particularly after day 15. The continuous increase in VCL, VAP, and VSL, with significant differences observed by day 30 and reaching peak values by day 90, can be attributed to the nutritional profile of the feed, which supports the improvement in sperm energy metabolism and motility mechanisms. The high levels of crude fat and protein in the WH-based diet (Table 2) likely contributed to the energy supply necessary for optimal sperm movement. These findings align with those in cattle and sheep, where nutrient-dense diets have led to improved sperm kinetics (Khademzade et al., 2022). The improvement in sperm kinematics over time may also indicate the beneficial effects of WH on mitochondrial function within sperm cells, as adequate nutrient intake supports mitochondrial energy production, which is essential for sustained motility and velocity (Srivastava et al., 2023).

Hormonal analysis revealed a marked increase in testosterone levels, which peaked on day 90, whereas FSH and LH levels remained relatively stable throughout the study. The nutritional content of the diet, particularly the influence of essential fatty acids and amino acids, which are precursors for testosterone synthesis, can explain the increase in testosterone (Gafer et al., 2023). Dietary interventions, particularly those involving plant-based feeds rich in omega-3 and omega-6 fatty acids, can stimulate testosterone production by influencing the hypothalamic-pituitary-gonadal axis, leading to increased secretion from Leydig cells (Ferramosca and Zara, 2022; Tabbsum et al., 2023). The hormonal responses were also notable. Testosterone levels progressively increased during the trial, whereas FSH and LH levels remained stable, indicating that the hypothalamic–pituitary–gonadal axis was not disrupted. The sustained increase in testosterone levels in this study could be a direct result of the optimized nutrient intake from the WH-based diet, which supports Leydig cell function and testosterone biosynthesis.

In contrast, the observed stable levels of FSH and LH suggest that the impact of WH feeding was more specific to testosterone production rather than broader gonadotropin regulation. This could be due to the fact that testosterone feedback mechanisms on the hypothalamus and pituitary are more sensitive than those regulating FSH and LH in rams. Previous studies on ruminants have similarly shown that FSH and LH are often less sensitive to dietary changes, although testosterone levels may fluctuate in response to nutrition (Abbara et al., 2024). The stability of gonadotropin levels may reflect hormonal homeostasis, while the increase in testosterone suggests enhanced Leydig cell function, which is potentially stimulated by the nutritional content of WH, which includes flavonoids and micronutrients known to support steroidogenesis. The hormonal profile in this study suggests that WH-based feeding may specifically support Leydig cell function, leading to improved testosterone synthesis without significantly altering FSH and LH concentrations (Santos et al., 2022).

The observed improvement in conception rates from day 45 onwards, with the highest values recorded at days 60 and 90, further supports the hypothesis that WH-based feeding enhances reproductive performance in Garut rams. The progressive increase in conception rates correlates well with improvements in sperm quality, motility, and hormonal regulation, indicating that the enhanced semen quality directly contributed to higher fertility. The increase in conception rates is consistent with findings from other studies, where dietary interventions leading to improvements in sperm motility and testosterone levels were also associated with higher fertility rates (Jurewicz et al., 2018; Salas-Huetos et al., 2019). The ability of WH-based diets to improve reproductive efficiency is promising, particularly in Garut rams, where fertility issues have been a limiting factor in breeding programs. These results emphasize the potential of using functional feed ingredients to optimize reproductive outcomes and increase productivity in sheep farming systems.

Although the study demonstrated promising results, several limitations should be acknowledged. The small sample size (10 rams) may limit the generalizability of the findings, and further studies with a larger animal cohort would strengthen the conclusions. In addition, the study did not explore the long-term effects of WH-based feeding on reproductive health and genetic performance. Future research should focus on the prolonged impact of this diet on fertility and its potential to influence offspring quality and genetic traits. Moreover, the role of environmental factors, such as temperature and humidity, which could influence reproductive outcomes and feed utilization, was not examined. These factors should be further investigated in subsequent studies to determine the optimal feeding conditions.

Correlation analyses revealed strong positive associations between semen parameters, hormone levels, and fertility outcomes, indicating a coherent physiological relationship influenced by the WH-based diet. Increased testosterone levels were positively correlated with improved sperm motility and progressive motility, suggesting that enhanced androgenic activity may play a key role in supporting spermatogenesis and overall semen quality. Additionally, sperm motility was significantly correlated with conception rates, highlighting the functional relevance of these improvements in reproductive success. These findings support the hypothesis that nutritional modulation through bioactive feed components can influence endocrine function and, in turn, enhance ejaculate quality and the likelihood of successful fertilization. Such correlations provide a mechanistic link between dietary intervention, reproductive physiology, and fertility performance, strengthening the biological plausibility of the observed outcomes.

The results of this study provide strong evidence for the potential of WH as a functional feed ingredient to improve the reproductive performance of Garut rams. WH-based diets could play a pivotal role in optimizing breeding efficiency by enhancing sperm quality, kinematics, and testosterone levels. The use of WH not only presents a sustainable approach to improving livestock productivity but also offers a viable solution for managing the overabundance of this invasive plant species in aquatic ecosystems. WH-based feeding strategies could contribute to the long-term sustainability of Garut ram breeding programs and provide economic benefits to local sheep farmers by improving reproductive health and conception rates. These findings highlight the importance of integrating functional feed ingredients into breeding programs to enhance productivity and environmental sustainability.

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Conclusion

In conclusion, this study demonstrates that WH-based diets can significantly enhance the reproductive performance of Garut rams, as evidenced by improvements in semen quality, sperm kinematics, hormonal regulation, and conception rates. WH’s nutritional profile, combined with its ability to positively influence testosterone production and sperm motility, offers a promising strategy for optimizing breeding programs in Garut rams. Moreover, the use of wh not only improves livestock productivity but also provides a sustainable solution to managing an invasive aquatic plant, contributing to both agricultural and environmental sustainability. These findings underscore the potential of wh-based feeding strategies to enhance the reproductive efficiency of sheep farming while promoting ecological conservation. Further research is required to explore the long-term effects and broader applications of this feeding approach.

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Acknowledgments

The authors would like to express their sincere gratitude to the Directorate of Research, Gadjah Mada University, for providing support through the 2025 Doctoral Research Grant (Kompetisi Doktor) program.

Conflict of Interest

The authors declare no conflict of interest.

Funding

This research was funded by the Doctoral Research Grant (Kompetisi Doktor) program of Universitas Gadjah Mada, grant no 1618/UN1/DITLIT/Dit-Lit/PT.01.00/2025. The authors gratefully acknowledge the support provided.

Authors’ contribution

AN, KDP, AMD, JBM, and KRP conceived and designed the experiments. AN, KDP, HH, and FAK performed the experimental procedures and laboratory analyses. HH and MSM supervised and coordinated the experiments and provided critical input during the study. AN and AMD performed the statistical analyses of the experimental data. AN, AMD, FAK, and MSM prepared the initial draft of the manuscript. All authors have critically revised the manuscript and approved the final version.

Data availability

All data are provided in the revised manuscript.

Abbreviations

AI, artificial insemination; CASA, computer-assisted sperm analysis; ELISA, enzyme-linked immunosorbent assay; FSH, follicle-stimulating hormone; LH, luteinizing hormone; VAP, average path velocity; VCL, curvilinear velocity; VSL, straight-line velocity.

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