Shahid Bahonar University of Kerman and Iranian Biotechnology Society Agricultural Biotechnology Journal 2228-6705 17 4 2025 10 23 Investigation of interleukin-6 gene expression in liver and spleen tissues of broiler chickens fed mealworm, probiotics, and mealworm with probiotics Investigation of interleukin-6 gene expression in liver and spleen tissues of broiler chickens fed mealworm, probiotics, and mealworm with probiotics 469 486 5155 10.22103/jab.2025.26228.1793 FA Mohammadreza Mohammadabadi Professor of Animal Science Department, Shahid Bahonar University of Kerman 0000-0002-1268-3043 Amin Khezri Animal Science Department, Faculty of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran. 0000-0002-5371-9831 Mohsen Afsharmanesh Animal Science Department, Faculty of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran. 0000-0002-4782-8687 Hamid Kheyrodin Semnan University, Semnan, Iran. Mohammad Reza Bahreini Behzadi Animal Science Department, Faculty of Agriculture, Yasouj University, Yasouj, Iran. 0000-0001-7092-3626 Olena Babenko Department of Animal Science, Bila Tserkva National Agrarian University, Bila Tserkva, Ukraine. 0000-0002-8404-3272 Svitlana Usenko Department of Animal Productivity Biology, Named after academician O.V. Kvasnytskyi, Poltava State Agrarian University, Poltava, Ukraine. 0000-0001-9263-5625 Mehdi Momen Department of Surgical Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, WI, USA. 0000-0002-2562-2741 Journal Article 2025 11 03 Objective Given the growing concerns about the use of antibiotics in the poultry industry and the search for natural alternatives to improve the performance and health of broiler chickens, and the fact that the interleukin-6 (IL-6) gene is a key indicator in the immune and inflammatory responses of birds, the present study was conducted to investigate the nutritional effects of mealworms (Tenebrio molitor) and probiotics, individually and in combination, on IL-6 gene expression in liver and spleen tissues of broiler chickens. Materials and Methods In this study, 160 one-day-old male chicks of the Ross 308 strain were used in a completely randomized design with four treatments and four replicates (each replication included 10 birds). The experimental treatments included the control group (basal diet), probiotic group (0.2 g/kg feed), mealworm powder group (0.5% of feed), and probiotic and mealworm combination group. The rearing period lasted 42 days. At the end of the period, tissue samples were taken from the liver and spleen for RNA extraction and cDNA synthesis. IL-6 gene expression was measured by Real-Time PCR using the GAPDH reference gene. Data were analyzed by ANOVA and Tukey's multiple comparison test (P<0.05). Results The results showed that the addition of probiotics, mealworm powder, and their combination led to a significant increase in IL-6 gene expression in liver and spleen tissues compared with the control group (P<0.05). The highest increase in expression was related to the combined probiotic and mealworm treatment, which was significantly different from other treatments. Single-peak melting curves and the correct size of PCR products confirmed the specificity of the reaction. Increased IL-6 expression in both tissues indicated stimulation of innate immune and acute phase inflammatory pathways at the systemic level. Conclusion The findings suggest that the combined use of probiotics and mealworm powder can have a synergistic effect on the immune activation of broiler chickens by increasing IL-6 gene expression. These results indicate that the combination of these two natural additives can be used as an effective and safe approach to promote immune health and sustainability of the poultry farming system. Objective Given the growing concerns about the use of antibiotics in the poultry industry and the search for natural alternatives to improve the performance and health of broiler chickens, and the fact that the interleukin-6 (IL-6) gene is a key indicator in the immune and inflammatory responses of birds, the present study was conducted to investigate the nutritional effects of mealworms (Tenebrio molitor) and probiotics, individually and in combination, on IL-6 gene expression in liver and spleen tissues of broiler chickens. Materials and Methods In this study, 160 one-day-old male chicks of the Ross 308 strain were used in a completely randomized design with four treatments and four replicates (each replication included 10 birds). The experimental treatments included the control group (basal diet), probiotic group (0.2 g/kg feed), mealworm powder group (0.5% of feed), and probiotic and mealworm combination group. The rearing period lasted 42 days. At the end of the period, tissue samples were taken from the liver and spleen for RNA extraction and cDNA synthesis. IL-6 gene expression was measured by Real-Time PCR using the GAPDH reference gene. Data were analyzed by ANOVA and Tukey's multiple comparison test (P<0.05). Results The results showed that the addition of probiotics, mealworm powder, and their combination led to a significant increase in IL-6 gene expression in liver and spleen tissues compared with the control group (P<0.05). The highest increase in expression was related to the combined probiotic and mealworm treatment, which was significantly different from other treatments. Single-peak melting curves and the correct size of PCR products confirmed the specificity of the reaction. Increased IL-6 expression in both tissues indicated stimulation of innate immune and acute phase inflammatory pathways at the systemic level. Conclusion The findings suggest that the combined use of probiotics and mealworm powder can have a synergistic effect on the immune activation of broiler chickens by increasing IL-6 gene expression. These results indicate that the combination of these two natural additives can be used as an effective and safe approach to promote immune health and sustainability of the poultry farming system.

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