Growth performance and immune response of broilers fed probiotic-supplemented diets

Introduction

How well a broiler flock converts feed into meat while staying healthy determines the profitability of any poultry enterprise. Over the past two decades, the removal of subtherapeutic antibiotics from broiler diets across much of the world has forced producers to look for safer growth-promoting alternatives [1] . Probiotics live microbial cultures that benefit the host when administered in adequate amounts have emerged as one of the most promising replacements [2] . Several species of Lactobacillus, Bacillus, and Saccharomyces have been tested individually or in combination in broiler feeding trials [3] . The proposed mechanisms through which probiotics act include competitive exclusion of enteric pathogens, production of short-chain fatty acids, modulation of the gut-associated lymphoid tissue, and improved nutrient digestibility [4] . However, the magnitude of growth response reported in the literature varies widely from negligible to a 12% improvement in final body weight depending on strain type, dose, bird age at first supplementation, and environmental stress level [5] . Immune modulation by probiotics is less well understood than growth effects. A few controlled trials have demonstrated elevated antibody titres against Newcastle disease virus in probiotic-fed broilers [6] , while others have recorded higher serum immunoglobulin concentrations [7] . But the relationship between dose level and immune response remains unclear, especially under tropical management conditions with high ambient temperatures and humidity [8] . In the Brazilian poultry sector, which produced roughly 14.8 million tonnes of chicken meat in 2022 [9] , cost-effective health strategies are in high demand. The objective of this research was to evaluate a commercial multi-strain probiotic at three graded inclusion levels on growth performance and humoral immune response in Cobb 500 broilers raised in a semicontrolled environment typical of Minas Gerais state.

Study area description

The experimental site lies in the Zona da Mata region of Minas Gerais at an elevation of 648 m above mean sea level. Viçosa municipality falls within the tropical highland climate zone (Köppen Cwb) and receives an average annual rainfall of 1,340 mm, most of which arrives between October and March. During the trial period (March-April 2023), ambient temperature outside the poultry house fluctuated between 22 °C and 32 °C, with relative humidity ranging from 58% to 85%. These warm-humid conditions are representative of commercial broiler production across much of southeastern Brazil and are known to impose moderate heat stress on rapidly growing Cobb strains.

Material and Methods

Material

This research was carried out at the experimental poultry unit of Federal University of Viçosa, Viçosa, Minas Gerais, Brazil (20.75°S, 42.88°W) from 10 March to 14 April 2023. Ethical clearance was obtained from the Institutional Animal Ethics Committee (Protocol No. CEUAP/UFV/2023-014, approved 18 February 2023). A total of 240 day-old Cobb 500 broiler chicks with an average hatching weight of 42.6 ± 1.3 g were procured from a local hatchery. Chicks were wing-banded, individually weighed, and randomly distributed into four treatment groups T0 (control), T1, T2, and T3 of 60 birds each, housed in three replicate pens of 20 birds. Floor pens measured 3 m × 2 m with rice husk litter at 5 cm depth. A standard corn-soyabean meal diet (22% CP starter, 20% CP finisher) was formulated to meet NRC (1994) recommendations [10] . The commercial probiotic (ProGut-Mix, containing Lactobacillus acidophilus, Bacillus subtilis, and Saccharomyces cerevisiae at 2 × 10⁹ CFU/g) was added to the basal diet at 0, 0.5, 1.0, and 1.5 g/kg for T0-T3 respectively.

Methods

Body weight and feed intake per pen were recorded weekly. Feed Conversion Ratio (FCR) was calculated as total feed consumed divided by total weight gain per replicate. Mortality was recorded daily. On day 35, blood samples (3 mL) were drawn from the brachial vein of six randomly selected birds per replicate (18 per treatment). Sera were separated by centrifugation at 3000 rpm for 15 min and stored at -20 °C. Newcastle Disease (ND) antibody titres were determined by Haemagglutination Inhibition (HI) test following standard OIE protocol [11] . Serum IgG and IgM concentrations were quantified using chicken-specific ELISA kits (Bethyl Laboratories, USA). Ambient temperature and relative humidity inside the house were logged twice daily using a digital thermo-hygrometer; the mean values during the trial were 28.6 ± 2.4 °C and 71.2 ± 6.1%, respectively. All data were subjected to one-way ANOVA using SPSS version 26.0. Treatment means were compared by Duncan's multiple range test at the 5% significance level [12] .

Results

Initial body weight at day 0 did not differ across treatments (p=0.87), confirming random allocation. By week 3, differences began to emerge, and at the end of the 35-day trial final body weights of T2 and T3 birds were significantly higher than those of T0 (p<0.05). Feed intake showed a modest but non-significant increase with probiotic supplementation. Immune parameters showed a clear dose-dependent trend.

ND HI titres at day 35 were significantly elevated in all three probiotic groups relative to the control (Table 2).

Serum IgG concentration rose progressively from T0 (4.21 mg/mL) to T3 (5.38 mg/mL), while IgM levels showed significant elevation only in T2 and T3 birds. The economic analysis revealed that despite the higher feed cost associated with probiotic inclusion, the improved FCR and heavier market weight in T2 offset the additional expense, yielding a 7.8% better return over feed cost compared with T0.

Comprehensive interpretation

Taken as a whole, these findings show that probiotic supplementation at 1.0 g/kg (T2) provided the best balance between growth improvement, immune enhancement, and cost efficiency. The 1.5 g/kg level (T3) offered only marginal further gains in body weight while increasing feed cost without a proportional economic return.

Discussion

The 11.2% improvement in final body weight observed in T2 birds agrees with the range (8-14%) reported by Kabir et al. [13] for Lactobacillus-based probiotics in tropical broiler operations. That the response became statistically detectable only from week 3 onward aligns with the time needed for probiotic organisms to colonise the gut mucosa and shift microbial ecology in favour of beneficial fermentation. A comparable lag was noted by Mountzouris et al. [14] in a European trial using a different multi-strain product.

The improved FCR in probiotic-fed groups likely stems from enhanced nutrient digestibility. Bacillus subtilis produces extracellular enzymes proteases, amylases, lipases that may complement the bird's own digestive secretions [15] . And the elevated short-chain fatty acid output associated with Lactobacillus fermentation lowers intestinal pH, creating an environment hostile to Clostridium and Salmonella but favourable for enterocyte proliferation.

Immunological data from this trial are consistent with the hypothesis that probiotics enhance humoral immunity through stimulation of gut-associated lymphoid tissue. The 19.7% increase in serum IgG seen in T2 birds mirrors findings by Haghighi et al. [7] who recorded a 15-22% rise following Lactobacillus administration in SPF chickens. ND antibody titres were highest in T3 but the difference between T2 and T3 was not significant, hinting at a plateau in immune stimulation beyond the 1.0 g/kg dose. One limitation of the present work is the relatively short duration (35 days). Longer trials extending to 42 or 49 days might reveal whether the growth advantage of probiotic groups widens or narrows over time.

Conclusion

This research set out to determine whether graded levels of a multi-strain probiotic could improve both growth and immune parameters in Cobb 500 broilers under warmhumid conditions. The data clearly show that supplementation at 1.0 g/kg feed increased final body weight by 11.2%, reduced FCR by 8.3%, and elevated serum IgG by 19.7% compared with the unsupplemented control. ND antibody titres improved across all probiotic groups, suggesting that even low-dose supplementation strengthens the humoral response to routine vaccination. For commercial producers in tropical regions seeking a costeffective alternative to antibiotic growth promoters, a probiotic dose of 1.0 g/kg appears to offer the strongest combination of performance and immune benefits. Future trials should examine the interaction of probiotic dose with heat-stress mitigation strategies such as dietary electrolyte balancing and ventilation management.

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