DEVELOPMENT OF INNOVATIVE INSECT MEALS AS FUNCTIONAL FEED INGREDIENTS FOR MONOGASTRIC ANIMALS

The global population is facing exponential growth, projected to exceed 9 billion by 2050, raising significant challenges for food security and sustainability. This rapid increase drives higher demand for food, particularly animal-based products, placing pressure on the agricultural and livestock sectors Addressing the resulting protein gap requires innovative, sustainable strategies that minimize the environmental impact of conventional feed sources like soybean and fishmeal. Sustainable animal nutrition has thus become a priority, focusing on optimizing resource efficiency, enhancing livestock health and productivity, and reducing antibiotic dependence. Key measures include incorporating alternative feed ingredients, by-products, and recycling nutrients from former feedstuffs. The One Health approach supports these efforts by integrating food safety, zoonotic disease control, and antimicrobial resistance management, recognizing the need for multidisciplinary strategies to support human, animal, and environmental health. In line with these challenges, the United Nations 2030 Agenda for Sustainable Development and the European Green Deal emphasize the importance of sustainable agricultural practices and the reduction of environmental impacts, while ensuring food security. In this context, insects offer a promising, sustainable protein source that can help reduce the reliance on arable land for animal feed crops production. By valorising food waste and agricultural by-products as insect substrates, this approach supports a circular economy, minimizing environmental impact and optimizing resource efficiency. Insect-derived meals offer a high-quality protein source combined with bioactive components like chitin and antimicrobial peptides. These components exhibit prebiotic effects, supporting beneficial bacterial populations and probably supporting intestinal integrity. Among various insect species, Tenebrio molitor (commonly known as mealworm) is notable for its valuable nutritional profile, primarily due to its high biological value proteins and functional components. This doctoral research aimed to advance the understanding of insect-based nutrition by exploring innovative rearing strategies for Tenebrio molitor larvae, with the goal of optimizing larval growth performance, enhancing the nutritional and functional profiles of insect-derived meals, and evaluating their application as a sustainable ingredient in pig nutrition. The first part of research focused on the evaluation of innovative hydration strategies (water with 12.7% of lactoferrin) and circular growth substrates from agro-industrial by-products (chestnut shell, carob, brewer’s spent grain), to optimize rearing methods and larvae performance. Obtained T. molitor larvae meals were analyzed for their nutritional composition (AOAC, 2023), main functional properties (antioxidant activity and inhibitory activity against E. coli strains) and metabolomic profile through nuclear magnetic resonance (NMR) spectroscopy. Obtained results showed that the substrate can influence the growth of T. molitor, as well as the morphological and chemical-functional properties of the resulting insect meals. Particularly, we observed that the inclusion of chestnut shell led to higher growth rate compared the control after 14 days, with values of 7% and 4%, respectively (p=0.0253). Similarly, carob and brewer’s spent grain included at 25% resulted in significantly enhanced growth rates of 12%, compared to 4% in the control (p=0.0395). Moreover, innovative rearing methods impacted the final color of insect meals. Specifically, 12.5% inclusion of chestnut shell showed a color variation (ΔE=5.59) compared to the control, as well as larvae hydrated with lactoferrin solution (ΔE =1.66). Regarding insect nutritional profile, differences in protein content were found, with increased levels registered in larvae reared on chestnut shell (12.5% inclusion) compared to the control, reporting values of 51% and 44%, respectively (p=0.0391). Additionally, variations in free amino acids and other metabolites were detected between chestnut-fed groups and the control suggesting that substrate composition can modulate insect metabolism. Protein increased was also observed in larvae hydrated with lactoferrin solution (46%; p=0.045). In terms of functional properties, larvae reared on chestnut shell and those hydrated with lactoferrin solution both exhibited higher antibacterial activity against E. coli strains compared to the control (p<0.05). These results highlight the potential of innovative rearing strategies to produce insect meals with enhanced nutritional and functional properties, promoting the sustainable use of agro-industrial by-products within a circular economy model. In the second part of this thesis, the attention was focalized on the expression of antimicrobial peptides (AMP) in larvae under different rearing condition, using real-time PCR, and on the bioactive role of chitin, major component of insect exoskeleton. Obtained results on AMP gene expression confirmed the presence of Tenecin 1, Tenecin 2, and Attacin 1a, which could play a positive role not only in supporting the insect’s immune system but also in enhancing the functional value of insect derived products as feed or food ingredients. The bioaccumulative capacity of insects could be explored to enhance their functional properties, as the inclusion of bioactive compounds in the rearing substrate may increase the antimicrobial potential of the resulting insect meals. The inclusion of bioactive compounds in the rearing substrate may influence insect metabolism, potentially modulating the antimicrobial properties of the resulting insect meals. In relation to the beneficial and safe role of insects in animal nutrition, the project also included an in-depth literature review on the prebiotic and anticancer effects of chitin. This analysis highlighted its potential to support gut health and contribute to animal health management strategies. Such review is particularly relevant in the context of the current infodemic, characterised by the spread of false news about the safety and health implications of insects as feed and food. As a functional ingredient, chitin aligns with the growing emphasis in animal nutrition on enhancing gastrointestinal function, which is closely linked to improved immune responses and to increased resistance to disease. The interaction between functional ingredients and gut microbiota can lead to improved strategies for enhancing gut health, which is essential for the overall welfare and disease resistance of animals. The third part of this research project aimed to investigate the use of T. molitor meals in pig diet on growth performance, nutrient digestibility, animal immune status, and the modulation of gut microbiota of pigs. With this purpose, a preliminary study was carried out on growing pig divided in two experimental groups receiving a basal complete diet (control, CON) and the same diet replacing 4% fermented soy protein concentrate with 5% T. molitor larvae meal. Findings demonstrated that the substitution of conventional proteins like soybean meal with T. molitor larvae meal can support animal performance without adverse effects. Moreover, T. molitor inclusion modulated gut microbiota composition, particularly at the genus level, with shifts in bacterial populations like Elusimicrobium and Asteroplasma, which are implicated in carbohydrate metabolism and antimicrobial activity, respectively. However, supplementary research is required to elucidate the functional implications of these changes and their impact on host physiology. In particular, a deeper understanding of the mechanisms driving microbiota modulation and gut health, coupled with efforts to address practical and regulatory challenges, will be essential to expand the inclusion of these innovative ingredients in livestock diets on a global scale. Future research should focus on determine optimal inclusion levels, characterizing bioactive compounds and exploring synergistic effects with other functional feed additives to maximize their benefits. In summary, insects represent a sustainable alternative protein source, requiring minimal land and water use. As efficient bio-converters, they transform low-nutritional value substrates into high-quality proteins, offering potential for optimizing production costs and promoting wider adoption in the animal feed sector. This approach supports the principles of the circular economy and contributes directly to the United Nations Sustainable Development Goals (SDGs), notably Goals 2 (Zero Hunger), 12 (Responsible Consumption and Production), and 13 (Climate Action). Additionally, the ability to rear insects in decentralized modular systems enhances their applicability, particularly in the swine industry, where cost-efficiency is a key consideration. The development of insect-based meals enriched with essential nutrients and antimicrobial peptides offers a promising strategy to address the global challenge of antibiotic resistance, providing a natural, integrable solution to improve animal health and reduce reliance on conventional antimicrobials.