Effects of fertilization and yeast culture supplementation on growth performance and nitrogen balance in Nellore young bullsABSTRACT.
We aimed to evaluate the effects of nitrogen (N) fertilization in tropical pastures and yeast culture (YC) supplementation on intake and digestibility, N use efficiency, and growth performance of Nellore young bulls during their growing phase. Treatments: 1) protein-energy supplementation at 0.3% body weight (BW) with YC (fertilized pasture 100 kg N ha-1 year-1), 2) protein-energy supplementation at 0.3% BW without YC (fertilized pasture 100 kg N-1 ha-1 year), 3) protein-energy supplementation at 0.3% BW with YC (unfertilized pasture), and 4) protein-energy supplementation at 0.3% BW without YC (unfertilized pasture). The experiment lasted 112 days and involved 60 Nellore young bulls (initial BW: 230 ± 10 kg) in a completely randomized design in a 2×2 factorial arrangement (Factor A: with/without fertilization; Factor B: with/without YC supplementation). Animals fed YC-supplemented diets consumed 145 g day-1 more crude protein, showed an additional average daily weight gain of 130 g day-1, and retained 20.4% more N (p<0.005) than animals fed diets without YC. Protein-energy supplementation at 0.3% of BW with YC addition effectively increased N retention in grazing Nellore young bulls and improved their growth performance. N fertilization in tropical pasture-based beef cattle systems, improved forage digestibility and animal production per unit area.
Keywords:
nitrogen fertilizer; xaraés grass; beef cattle; growing phase; feed additive
Introduction
Increasing productivity in beef cattle production has become crucial in recent years, as the failure to adopt modern technologies makes the system less economically competitive and sustainable. Strategies such as N fertilization, pasture management, and dietary supplementation with additives have been employed to increase production efficiency and sustainability (Cardoso et al., 2016; Teobaldo et al., 2023).
N fertilization practices and supplementation increase N retention, reduce urinary excretion and, consequently, decrease nitrous oxide emissions (Cardoso et al., 2020; Dallantonia et al., 2021; D’Aurea et al., 2021; Fonseca et al., 2022). Strategic supplementation can result in additional daily weight gains of approximately 0.250 kg animal-1 day during the rainy season (Koscheck et al., 2020; Berça et al., 2023; Teobaldo et al., 2023; Fonseca et al., 2024).
Among the various feed additives used in protein-energy supplements, prebiotics have emerged as a promising alternative to antibiotics (e.g., probiotics such as virginiamycin and monensin), which have raised increasing concerns in recent years due to the risks associated with antibiotic use in animal feed. These prebiotic additives serve as strategies to reduce the duration of the growing phase, increase forage intake, and significantly increase animal weight gain (Lambo et al., 2021).
Saccharomyces cerevisiae yeast culture (YC) is an additive consisting of inactive yeast biomass and controlled fermentation metabolites (Shurson, 2018). Feed additives, such as YC, are used to improve production system efficiency by stimulating the growth of beneficial ruminal microorganisms, including bacteria that ferment fibrous carbohydrates, thereby improving ruminal fiber digestion (Amin & Mao, 2021; Kholif et al., 2024).
The increase in cellulolytic bacteria populations due to microbial growth leads to higher ammonia N requirements by bacteria. Thus, inadequate ammonia N supply may lead to decreased ruminal concentrations. Therefore, N supplementation is essential to increase microbial protein synthesis efficiency and, consequently, improve N use efficiency (NUE) (Adesogan et al., 2018).
Based on these considerations, we tested the following hypotheses: 1) dietary YC supplementation improves the growth performance of animals; 2) YC intake reduces urinary N excretion; 3) YC improves dry matter (DM) digestibility in unfertilized pastures; and 4) N fertilization increases weight gain per unit area.
Therefore, in this study, we evaluated the effects of protein-energy supplementation at 0.3% of body weight (BW), with or without YC, on DM and nutrient intake and digestibility, NUE, and the growth performance of Nellore young bulls grazing on Xaraés grass pastures, with or without N fertilization.
Materials and methods
All procedures followed the Ethical Principles in Animal Research adopted by the National Council for Animal Experimentation Control and were approved by the Institutional Animal Care and Use Committee of São Paulo State University (UNESP), Jaboticabal Campus, under protocol 254/22). The study was conducted at the Forage Research Unit of the School of Agricultural and Veterinary Sciences, ‘Júlio de Mesquita Filho’ UNESP, in Jaboticabal, São Paulo State, Brazil. The study site is located at 21º14′05′′ S, 48º17′09′′ W, at an elevation of 615 m.
Animal evaluations were conducted on a 22-ha experimental area established with Urochloa brizantha (syn. Brachiaria brizantha) (Hochst. ex A. Rich.) Stapf cv. ‘Xaraés,’ subdivided into 12 experimental paddocks and a 0.5-ha reserve area.
Experiment, animals, and grazing method
The experiment was conducted during the animal growing phase over 112 days, divided into four 28-day periods, from December 2021 to April 2022. In total, 120 Nellore (Bos taurus indicus) young bulls with an initial BW of 230 ± 10 kg and age of 12 ± 2 months were used. The grazing method employed continuous stocking with variable stocking rates, involving 120 animals, 60 as testers and 60 as regulators, following the ‘put and take’ technique, to maintain a sward height of 30 cm, corresponding to 95% light interception (LI).
Treatments and experimental design
The 60 tester animals were randomly allocated to four treatments with three replicates (paddocks) each, totaling 12 experimental units, in a 2 × 2 factorial arrangement (A × B; Factor A: fertilized and unfertilized pasture, and Factor B: with and without YC supplementation). The treatments were as follows: Treatment 1 ‒ protein-energy supplement at 0.3% of BW with YC inclusion (YC + pasture fertilized with 100 kg N ha-1 year-1); Treatment 2 ‒ protein-energy supplement (control) at 0.3% of BW (pasture fertilized with 100 kg N ha-1 year-1); Treatment 3 ‒ protein-energy supplement at 0.3% of BW with YC inclusion (YC + unfertilized pasture); and Treatment 4 ‒ protein-energy supplement (control) at 0.3% of BW (unfertilized pasture). The YC of S. cerevisiae (Cultron® Aleris Nutrition; Jundiaí, São Paulo, Brazil) was used at a proportion of 1.2% of the supplement (Table 1) to ensure a minimum intake of 7 g animal-1 day-1 through supplementation.
The protein-energy supplements, consisting of ground corn and soybean meal (Table 1), were provided daily at 9h. Before the experimental period, animals underwent a 15-day adaptation phase with gradual supplementation enhancement every 5 days from 0.1 to 0.2% of BW and then to 0.3% of BW.
The paddocks (n = 6) received split-application N fertilization (50 kg N ha-1 year-1 application) broadcast twice during the experimental period, totaling 100 kg N ha-1 year, with applications on January 18 and March 8, 2022.
Forage mass, morphological components, and chemical composition
Canopy height was measured weekly using a sward stick at 80 random points per hectare to adjust the stocking rate and maintain a sward height of 30 cm. Forage mass (FM) was estimated every 28 days by cutting and collecting four forage samples from representative points (average height) in each paddock.
The forage was harvested at ground level using a 0.25 m2 circular ring. Samples were initially weighed and divided into two subsamples. One subsample was used to determine the morphological composition of the pastures, manually separating dead material (leaves and stems), green stems, and green leaves. The other subsample was used to estimate the total DM availability (kg DM-1 ha-1) of forage in each paddock (Table 2). Samples were oven-dried at 55°C for 72h in a forced-air circulation system and then weighed.
To determine the chemical composition of the forage, pasture samples representing the fraction consumed by the animals were collected every 28 days using the hand-plucking method (Halls, 1954), which involves harvesting forage after observing the grazing behavior of animals. Fresh samples were oven-dried at 55°C for 72h under forced air circulation. Following pre-drying, all forage samples were ground to 1 mm and analyzed for DM, organic matter (OM), ether extract, crude protein (CP), and fiber fraction components using near-infrared spectroscopy (NIRS) with a Fourier transform near-infrared (FT-NIR) modular spectrometer (NIRFlex® N500; Büchi Labortechnik AG, Flawil, Switzerland).
Intake and digestibility
Nutrient intake and digestibility were evaluated in 9 tester animals per treatment (3 animals/ paddock, totaling 36 animals) after completing their growth performance assessment, with evaluations conducted between days 90 and 100 of the experimental period.
Estimates were obtained using indigestible neutral detergent fiber (iNDF) as an internal marker based on fecal output data. To estimate fecal output, chromic oxide was administered daily (10 g animal-1) in paper capsules for 10 consecutive days at the same time of the day, with a 7-day adaptation period followed by a 3-day fecal collection period (Hopper et al., 1978).
Fecal samples were collected once daily at different times as follows: day 1 at 16h, day 2 at 11h, and day 3 at 6h. The collected samples were oven-dried at 55°C for 72h in a forced-air circulation system.
Titanium dioxide (TiO2) was used as an external marker to estimate individual supplement intake (Titgemeyer et al., 2001). Animals were administered 10 g day-1 of TiO2, mixed into the supplement immediately before feeding, for a duration of 10 days.
Nutrient digestibility was estimated by quantifying the iNDF content in forage samples collected through simulated grazing, feces, and protein-energy supplement using NIRS with an FT-NIR modular spectrometer (NIRFlex® N500; Büchi Labortechnik AG, Switzerland).
NUE and microbial protein synthesis
Urine samples were collected by spontaneous micturition between days 90 and 100 of the experimental period. Nine tester animals per treatment (3 animals/paddock; totaling 36 animals) were evaluated after they were assessed for their growth performance. To estimate urinary volume, N concentration, and other urinary compounds, spot urine samples (50 mL) were collected from each animal following the methodology of Chizzotti et al. (2008). Urine samples were immediately filtered and diluted by mixing their 10 mL aliquots with 40 mL of 0.072 M sulfuric acid to prevent the bacterial degradation of purine derivatives and uric acid precipitation.
Purine derivative (PD) concentrations, including uric acid and allantoin, were determined using Analisa® commercial kits through enzymatic-colorimetric methodology and the Fujihara et al. (1987) methodology, as outlined by Chen and Gomes (1992). Absorbed purines were calculated based on PD excretion (Orellana Boero et al., 2001).
The ruminal synthesis of nitrogenous compounds was estimated based on absorbed purines (Chen & Gomes, 1992). Thus, NUE was calculated as the ratio between N balance (N intake minus N excretion in feces and urine) and total N intake from pasture and supplement (Detmann et al., 2014). Another aliquot of concentrated composite urine was used to determine total N content using the Kjeldahl method (Fenner, 1965) and through urea concentration using Analisa® commercial kits via enzymatic colorimetric analysis.
Urinary creatinine concentration in the spot sample was measured by colorimetric analysis using Analisa® commercial kits with alkaline picrate, and the obtained value was used to estimate urine volume. Urinary N excretion was calculated by multiplying urinary volume by urinary N concentration (Chizzotti et al., 2008).
Growth performance of animals
Average daily weight gain (ADG) was calculated by weighing the animals on days 0 and 84 of the experimental period, following a 14h feed and water fasting. Additionally, animals were weighed every 28 days without fasting to adjust stocking rates, monitor health status, and administer deworming treatments and mandatory vaccinations as required.
Weight gain per unit area was calculated based on the average individual weight gains of tester animals and the number of animals per paddock during the evaluation period accounting for the number of animal-days. Data for the animals used as regulators were included in the calculations of stocking rate and weight gain per unit area, accounting for their occupation days in each paddock. The stocking rate, expressed in animal units (AUs) per hectare (1 AU = 450 kg BW ha-1), was determined based on the total BW of animals maintained in each paddock during each experimental period.
Statistical analyses
Performance, intake, digestibility, and N balance data were analyzed as a completely randomized design in a 2 × 2 factorial arrangement (with or without fertilization × with or without YC) using the SAS PROC MIXED software (SAS Institute, 2008 ). The average measurements of five animals (growth performance) and three animals (intake, digestibility, and balance) per paddock were analyzed as a single value (paddock as the experimental unit). Statistical analyses were conducted with four treatments and three replicates per treatment (n = 12). The normality of errors and the homoscedasticity of data were verified using PROC UNIVARIATE in SAS® (SAS Institute, 2008). The models included the main effects of fertilization (with or without), YC supplementation (with or without), and their interaction (fertilization × YC). Treatments were compared using Tukey’s test (p<0.05) in the SAS PROC MIXED software (SAS Institute, 2008).
Results and discussion
Intake and digestibility
There was no interaction between N fertilization and YC supplementation on intake and digestibility data (p>0.05). Neither N fertilization nor YC supplementation affected DM intake, OM intake, or total digestible nutrients (TDN) (p>0.05). However, the intake of CP and neutral detergent fiber (NDF) corrected for ash and protein (NDFap) were higher in animals fed diets supplemented with YC (p<0.05; Table 3). Although YC supplementation did not impact DM or nutrient digestibility (p>0.05), animals grazing on N-fertilized pastures exhibited higher digestibility values for DM, OM, CP, and NDFap (p<0.05; Table 3).
According to Reis et al. (2009), forage intake is impacted by nutritional and non-nutritional factors, wherein nutritional factors are determined by the basal diet. In particular, insoluble fiber plays a particularly significant role by directly contributing to the effects of rumen fill, thereby regulating intake.
However, non-nutritional factors are linked to the canopy structure available for intake and forage selection, rate of intake (bite size × rate), and grazing time, with the adopted pasture management resulting in similar effects of these variables on intake (Reis et al., 2009).
Adjusting the stocking rate to maintain a canopy height of 30 cm resulted in similar forage supply across all treatments, leading to similar nutrient intake, except for CP and NDFap, which were impacted by the nutritive value of forage and dietary supplements. The increase in the mean intake of CP and NDFap in response to dietary supplementation is likely because of the composition of the dietary supplement containing YC (Table 3).
N-fertilized pastures subjected to continuous stocking and managed with stocking rate adjustments based on pasture management criteria, such as 95% LI, height, herbage mass, or forage supply, enable continuous consumption of young leaves by grazing animals, which results in high digestibility of consumed forage (Hoffmann et al., 2021; Leite et al., 2021; Berça et al., 2023; Quadros et al., 2023; Fonseca et al., 2024). In the present study, higher digestibility values were observed for DM (mean 58%), CP (mean 55%), and NDFap (mean 59%) when pastures received N fertilization (Table 3). This can be attributed to the chemical composition of the consumed forage, with a higher concentration of dead material (7% higher) observed in pastures that did not receive N fertilization (Table 2).
Erasmus et al. (1992) reported improved digestibility coefficients for DM, CP, and fiber in dairy cows fed diets supplemented with yeast culture. Gunun et al. (2022) reported improved CP digestibility in cattle fed with diets supplemented with yeast at the concentrations of 0, 1, 2, and 3 g kg-1 DM. In the present study, animals consumed 1.4 g kg-1 DM of YC without any effect on digestibility coefficients, consistent with the findings of Gomes et al. (2011), who observed no improvements in digestibility coefficients when 0.6 g kg-1 DM of S. cerevisiae was included in the diets of Nellore steers. Similarly, Garcia et al. (2000) reported no differences in nutrient digestibility in zebu cattle fed with diets supplemented with yeast. These variable responses may be attributed to factors such as supplementation levels, differences in yeast products, dietary composition, or animal-specific characteristics (Hansen et al., 2017; Zhang et al., 2022).
NUE and microbial protein synthesis
No interaction was observed between the effects of fertilization and YC supplementation. N fertilization did not affect N utilization and microbial protein synthesis (p>0.05). However, animals supplemented with YC showed higher N intake (p=0.007), greater fecal N excretion (p=0.014), and greater N retention compared to animals that did not receive the additive (p=0.009; Table 4).
Studies indicate that regardless of ruminant species or category, fecal and urinary N excretion increases with higher N intake (Schuba et al., 2017). In the present study, animals receiving YC consumed an average of 23 g day-1 more N and exhibited an average of 12 g day-1 higher fecal N than that in the animals in the control group (Table 4), which is consistent with the results reported in earlier studies (Schuba et al., 2017).
Vandehaar and St-Pierre (2006) stated that fecal N excretion is influenced by undigested ruminal N, undigested microbial protein in the small intestine, and endogenous N sources. Thus, Costa-Roura et al. (2020) reported that reducing N loss through urine or feces can be achieved by decreasing dietary N levels and protein degradation, as well as by improving the capture efficiency of rumen-degraded N through microbial protein synthesis, which may explain the lack of difference in urinary N excretion (Table 4).
Martins et al. (2023) supplemented 14 g cow-1 day-1 of YC and reported reduced urinary N excretion, indicating the potential for improved N utilization. Liu et al. (2022) reported that supplementing 2 and 4 g head-1 day-1 of YC can improve N utilization and reduce the environmental impact on beef cattle production. Ezequiel et al. (2000) reported that yeast supplementation improved N utilization, as observed in the present study where animals consuming 1.4 g-1 kg-1 DM of YC retained 20.4% more N than in animals fed diets without YC supplementation (Table 4).
N retention refers to its utilization in tissue protein synthesis, which can be directed toward new tissue formation, enzymatic systems, tissue replacement, or epithelial maintenance. The efficiency of N compound utilization by animals depends on the quality of these compounds reaching the tissues after intestinal absorption (Silva et al., 2019). The dietary protein provided by YC supplementation was likely more efficiently utilized in the post-ruminal environment, contributing to an increase in metabolizable protein content.
The efficiency of microbial protein synthesis relies on the synchronized availability of fermentable carbohydrates and N in the rumen, reaching its maximum when carbohydrate and protein degradation occur simultaneously (Santos et al., 2021). The absence of differences across treatments indicates that neither YC supplementation nor N fertilization of the pasture significantly improved ruminal microbial protein synthesis. Notably, well-managed tropical pastures naturally promote efficient microbial protein synthesis because of their high soluble N content, high-quality fiber, and low iNDF content.
Poppi and McLennan (1995) indicated that maximum efficiency of microbial protein synthesis and dietary protein transfer to the intestine occurs at levels below 160 g CP kg-1 of DOM, whereas levels exceeding 210 g CP kg-1 of DOM result in reduced transfer of CP from intake to the small intestine. However, intensively fertilized and managed tropical pastures have values higher than 210 g BW kg/DOM (Fonseca et al., 2022; 2024), as observed in our study, at approximately 271 g BW kg/DOM.
Growth performance of animals
No interaction was observed between the effects of N fertilization and YC supplementation. Although N fertilization did not impact ADG (p = 0.876), it significantly increased stocking rates by approximately 0.5 AU/ha (p = 0.0001). YC supplementation, however, improved ADG by 130 grams day-1 compared to the control group (p = 0.005; Table 5).
ADG responds to the supply of supplementary nutrients through supplementation (Sollenberger & Vanzant et al., 2011). The ADG of animals receiving protein-energy supplementation (0.3% of BW) with YC inclusion was 130 g day-1 higher than in animals fed diets without yeast supplementation, likely because of increased CP intake (Table 3).
Oliveira et al. (2023) investigated the effects of supplementation of 7 g animal-1 day-1 of yeast in feedlot cattle diets and observed improved final BW gain, attributed to enhanced diet digestibility. Although YC supplementation did not directly affect energy intake and digestibility, its impact on ruminal fermentation likely contributed to increased individual weight gain observed in animals receiving YC. Similarly, Wagner et al. (2016), in their review, highlighted that YC increases the production of propionic acid, a gluconeogenic volatile fatty acid, which may explain the improved N retention observed in animals fed YC-supplemented diet (Table 4), ultimately contributing to the positive effect on ADG (Table 3).
Peng et al. (2020) reported that supplementing beef cattle with 2 g animal-1 day-1 of yeast reduced inflammatory parameters, potentially leading to improved growth performance of animals.
Fertilized pastures led to increased stocking rates (AU/ha) and weight gains per unit area (kg ha-1). N fertilization primarily results in increased FM production (Delevatti et al., 2019), leading to higher stocking rates and, consequently, improved weight gain per unit area, as shown in Table 5.
The average Brazilian stocking rate is 0.94 AU ha-1 (Associação Brasileira das Indústrias Exportadoras de Carne, 2023), which is considerably lower than the results of this study. Here, N fertilization achieved an average stocking rate of 3.2 AU ha-1, compared to 2.4 AU ha-1 in unfertilized pastures. These findings highlight the exceptional quality of Xaraés grass pastures under continuous grazing with variable stocking rate when maintained at a 30-cm height. Moreover, protein-energy supplementation (0.3% of BW) facilitated increased grazing intensity, leading to higher stocking rates and, consequently, greater weight gain per unit area.
Conclusion
Our findings provide valuable insights for developing effective strategies to improve nitrogen balance and growth performance in Nellore young bulls. N fertilization in beef cattle production systems on tropical pastures during the rainy season is a well-established strategy to enhance forage digestibility and boost animal productivity per unit area.
The dietary administration of the YC-containing protein-energy supplement at 0.3% of BW to Nellore young bulls grazing on Xaraés grass during the rainy season effectively improved their ADG and overall growth performance, which can be attributed to increased CP intake and improved N retention in the animals.
Data availability
Does not apply.
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