Partial to complete replacement of soybean meal with high-protein wheat in broiler diets: Effects on growth and blood parameters

2.1. Animal ethics

All experimental procedures, including the humane euthanasia of birds for blood sampling via CO₂ stunning and decapitation, were reviewed and approved by the Animal Welfare Advisory Team at Wrocław University of Environmental and Life Sciences (Ethical statement no. 04/2023) in compliance with Directive 2010/63/EU.

2.2. Feed component

The study evaluated HPW supplied by Saatbau Poland Sp. z o.o. (Środa Śląska, Poland) as a protein substitute in broiler diets.

The HPW utilized in this study was Activus, a winter wheat cultivar classified in the A quality class, bred by Saatbau Linz GmbH (Austria). This cultivar is characterized by an elevated grain protein content and high baking quality. The batch used in this trial was analysed for its basic nutritional profile, showing a total protein content of 16% (Szuba-Trznadel et al., 2024), making it suitable for high-spec poultry diet formulations.

2.3. Animals and husbandry

The experiment was conducted at the Research and Education Station in Swojczyce, Wrocław University of Environmental and Life Sciences. A total of 196 one-day-old Ross 308 broiler chickens were obtained from ModerHatch hatchery (Magnuszowice, Poland). Upon arrival, broilers were individually weighed and randomly allocated to pens to achieve a uniform initial pen body weight (BW) (46 g ± 1.53 per pen), following a completely randomized design.

Birds were assigned to four experimental groups: group I (control), and groups II, III, and IV. Each group consisted of seven pens with seven birds per pen. Broilers were reared under standard conditions in accordance with Polish animal welfare regulations (Journal of Laws of the Republic of Poland, 2015, item 266, as amended in 2023, item 465) and Ross 308 management guidelines (Ross Broiler Management Handbook, 2018).

The ambient temperature was set at 32°C on day 1, gradually reduced to 24°C by the end of the second week, and maintained at 20–21°C thereafter. Relative humidity ranged from 60 to 73%. The lighting program provided 22 h of light and 2 h of darkness during the first week, adjusted to 18 h of light and 6 h of darkness from the second week onward. Feed and water were provided ad libitum. Each pen was equipped with a hopper feeder and two nipple drinkers with adjustable heights.

2.4. Dietary treatments

Birds were fed starter (days 1–14), grower (days 15–29), and finisher (days 30–49) diets. The finisher phase was extended by 1 week beyond the typical rearing period. Group I served as a wheat-based control (58% inclusion), while groups II–IV received diets with progressively increasing levels of HPW at the expense of SBM. The detailed composition of experimental diets is shown in Tables 1–3. Changes in SBM and HPW inclusion across feeding phases are presented in Table 4, while selected nutritional parameters are shown in Table 5.

All diets were formulated to be isoenergetic. ME was standardized at approximately 12.68 MJ·ME·kg⁻¹ (starter), 13.02 MJ ME·kg⁻¹ (grower), and 13.18 MJ ME·kg⁻¹ (finisher) by adjusting soybean oil inclusion. While the CP content in group I was slightly higher than Polish standard recommendations, the formulation was adjusted in collaboration with the feed manufacturer (Neorol Sp. z o.o.) to align with the specific nutritional requirements of the Ross 308 lineage. Conversely, in groups II–IV, the CP content was intentionally and progressively reduced below reference values as HPW inclusion increased. EAAs requirements were met via crystalline amino acid supplementation according to Polish Nutritional Recommendations for Poultry (Smulikowska & Rutkowski, 2018). To mitigate potential adverse effects of high HPW inclusion, all diets were supplemented with commercial enzyme preparations: xylanase (Ronozyme^® WX, DSM Nutritional Products, The Netherlands), containing endo–1,4-β-xylanase (EC 3.2.1.8); guaranteed minimum activity of 1,000 FXU·g⁻¹, included at 0.02%, providing a final activity of 200 FXU·kg⁻¹ of feed and phytase (Ronozyme^® HiPhos, DSM Nutritional Products, The Netherlands), containing 6-phytase (EC 3.1.3.26); guaranteed minimum activity of 10,000 FYT·g⁻¹, included at 0.02%, providing a final activity of 2,000 FYT·kg⁻¹ of feed.

2.5. Performance measurement and sampling

The experimental period was extended to 49 days based on the specific nature of the dietary treatments (high to complete replacement of SBM with HPW). This extension enabled the assessment of altered growth trajectories and compensatory growth, as well as the physiological maturity and quality of the carcass.

2.6. Data analysis

Data were organized using Microsoft Excel (Microsoft Corp., Redmond, WA, USA, 2019). Statistical analyses were performed with Statistica 13 (Tibco Software Inc., Palo Alto, CA, USA, 2017). Data distribution was checked separately within each group using the Shapiro–Wilk test. Homogeneity of variance between groups was assessed with Levene’s test. One-way analysis of variance (ANOVA) was used for statistical comparison, followed by Tukey’s honestly significant difference test to determine significant differences between group means. Statistical significance was set at P < 0.05 and P < 0.01. Obtained results are presented in tables and figures. Results in tables are presented as arithmetic means with the standard error of the mean (SEM). The SEM values presented in the tables represent pooled SEMs calculated from the residual mean square error obtained from the ANOVA/generalized linear model. Figures 1 and 2 show means with error bars. Significant differences between groups (indicated by different superscript letters) and corresponding P-values are presented in tables below each figure.

The line chart shows weekly body weight gain in broiler chickens from days 7 to 49 in four dietary groups, alongside the Ross 308 reference values. BWG generally increased with age, although it fluctuated between weeks. Group 4 consistently showed the lowest BWG. Group 3 also generally had lower values than Group 1 and 2. Group 2 exceeded Group 1 during days 35–42 and had a similar value during the final week. The Ross 308 reference increased to a peak around day 42 and then declined slightly. Error bars represent SEM; statistical differences are indicated in the accompanying table.

Figure 1

The line chart presents average daily feed intake across seven weekly intervals, from days 1–7 to days 42–49, for four dietary groups and the ross 308 reference. Feed intake generally increased with age. Group 4 had the lowest intake in every interval, with particularly large differences during the early weeks. Group 3 had lower intake than Groups 1 and 2 during days 28–35. Intake in Groups 1–3 was more similar during the later weeks, while Group 4 remained below the other groups and the reference. Error bars represent SEM; statistical differences are indicated in the accompanying table.

Figure 2

3.1. Performance

Weekly BW values of broiler chickens are presented in Table 6. No significant differences among groups were observed during the first week of life. However, from day 7 onward, birds in group I (58% HPW) and group II (71.1% HPW) exhibited significantly higher BW (P < 0.01) compared with groups III (77.6% HPW) and IV (84.1% HPW). On day 14, groups I and II remained heavier (P < 0.01) than group IV, while group III showed a significant reduction in BW compared to both the control group and group II (P < 0.05).

By day 21, BW decreased linearly as HPW inclusion increased (P < 0.01), with the highest BW recorded in the control group. This linear pattern persisted through days 28 and 35 (P < 0.01). By day 42, no significant difference was observed between groups I and II; however, both were significantly heavier than groups III and IV (P < 0.01). At the end of the experiment (day 49), groups I and II exhibited comparable BW, whereas groups III and IV remained significantly lighter (P < 0.01). Overall, group II maintained a BW similar to the control group, while higher HPW inclusion levels resulted in BW reductions of approximately 16% and 42% in groups III and IV, respectively.

Daily BWG is illustrated in Figure 1. Notably, group II showed an upward trend in BWG during the later stages of rearing, approaching or even exceeding control values.

FI across the rearing phases is summarized in Figure 2. During the starter phase, group IV consumed significantly less feed (14.85 g·day^–1·bird^–1) than the control group (20.35 g·day^–1·bird^–1; P < 0.01). Between days 7 and 14, FI was comparable among groups with HPW inclusion up to 77.6% (64.29–67.96 g·day^–1·bird^–1) but remained significantly lower in group IV (38.28 g·day^–1·bird^–1; P < 0.01). This reduction in FI associated with higher HPW inclusion persisted throughout the duration of the experiment.

FCR results are presented in Table 7. During the first week, FCR averaged 0.75 kg·kg^–1, with no significant differences among groups. From the second week onward, group IV consistently exhibited the poorest (highest) FCR (P < 0.05 to P < 0.01), whereas groups I and II remained comparable. This trend persisted throughout the entire experimental period.

Regarding mortality, no bird losses or health complications were observed during the entire 49-day experimental period. Because there was no mortality, no adjustments for missing bird-days due to the death were required in the final FCR and EPEF calculations.

3.2. Rearing efficiency indicators

3.3. Blood parameters

The effects of dietary HPW inclusion on selected haematological and biochemical blood parameters are presented in Tables 12–15.

4.1. Production performance

The progressive reduction in bird numbers per pen during the experiment may have influenced FI and contributed to increased variability in performance parameters, particularly during the final phase. Previous studies have shown that a reduction in stocking density below certain thresholds can affect FI and growth performance. Moreover, it has been reported that low stocking density can increase the coefficient of variation in BW (Feddes et al., 2002). While this reduction was a necessary consequence of the longitudinal sampling design, it should be considered when interpreting the finisher phase data.

In the present study, high inclusion levels of HPW, particularly above 77%, were associated with reduced BW. The pronounced reduction in BW observed in group IV (–42% vs. group I at day 49) appears to be driven primarily by markedly reduced FI, which directly limited the energy and amino acid supply necessary for growth. This is supported by the substantially lower FI recorded in this group, suggesting that reduced nutrient intake, rather than the diet composition per se, was the primary constraint on performance. A strong relationship between FI and growth rate in broilers has been consistently demonstrated in previous studies (Selle et al., 2023). In addition, several secondary mechanisms may have contributed to this growth depression. High inclusion levels of wheat are associated with increased NSP content, which may elevate digesta viscosity and impair nutrient digestibility (Selle et al., 2023). Although xylanase was included in the present study, its efficacy depends on the enzyme dose, substrate characteristics, and gut conditions. Consequently, it may not have been sufficient to fully counteract NSP-related effects at such extreme wheat inclusion levels (84%). Therefore, the growth failure in group IV is best characterized as a multi-factorial response, in which reduced FI acted as the primary driver, while NSP-related effects on digestibility likely contributed additively. This interpretation aligns with previous reports, indicating that excessive wheat inclusion, particularly under reduced CP conditions, can impair performance through the combined effects of diminished intake, impaired digestibility, and reduced metabolic efficiency (Choct et al., 2004; Selle et al., 2023). Comparable results were reported by Steenfeldt (2001), who observed decreased BW in broilers fed diets containing 81.5% wheat derived from 16 cultivars (112–127 g·kg^–1 CP), compared with diets containing 65% wheat. Choct et al. (2004) reported improvements in daily BWG, FCR, and excreta moisture content in broilers fed xylanase-supplemented wheat-based diets under low-energy conditions, suggesting that enzyme supplementation may partially alleviate the negative effects of high wheat inclusion. Furthermore, it has also been suggested that higher xylanase inclusion rates, the incorporation of 1–3% insoluble fibre, or the use of coarser wheat particle size may further mitigate the adverse effects of arabinoxylans on gut function (Hikawczuk et al., 2023; Naeem et al., 2024).

The absence of significant differences in BW between group I (control) and group II during both the starter (71.1% HPW) and grower (77.8% HPW) phases, combined with the relatively high CP content of the HPW used (163 g·kg^–1), indicates that partial replacement of SBM with HPW is feasible during early rearing. The lack of significant differences in FCR among groups during the first week of life is likely attributable to residual yolk sac reserves providing a buffer for nutritional requirements.

Biesek et al. (2022) demonstrated that partial substitution of ground wheat with whole wheat grain (10% of the diet) improved BW and FI without affecting FCR. Similarly, Yin et al. (2020) observed no significant differences in BW and FI between broilers fed high CP (215 g·kg^–1) and low-CP (165 g·kg^–1) wheat-based diets, although FCR was slightly higher in the low-CP group (1.58 vs. 1.49 kg·kg^–1). Interestingly, mortality was numerically lower in birds fed whole wheat, without detrimental effects on performance.

Reducing CP levels in wheat-based broiler diets remains a significant challenge, as it is frequently associated with impaired growth performance, characterized by reduced BWG and increased FCR (Selle et al., 2023). In the present study, feed efficiency during later rearing phases was most favourable in groups I and II, whereas higher HPW inclusion levels (groups III and IV) were associated with increased FI per kilogram of BWG. Similar findings were reported by Wang et al. (2024), who observed increased FI and FCR when wheat inclusion reached 55.77% during the grower phase and 62.38% during the finisher phase, despite enzyme supplementation. Chrystal et al. (2021) reported that reducing dietary CP to 193 g·kg^–1 in wheat-based diets (63.7% wheat inclusion) between days 7–35 resulted in only a 0.42% reduction in daily BWG and a 3.9% increase in FCR compared with maize-based diets. Likewise, Greenhalgh et al. (2022) reported a 3.39% reduction in BWG and a 3.47% increase in FCR when dietary CP was reduced to 180 g·kg^–1 in wheat-based diets during days 21–35.

4.2. Rearing efficiency indicators