INTRODUCTION

Projections indicate that the global egg production industry will experience a 39% increase in demand between 2005 and 2030. This expansion has been accompanied by increasing consumer expectations regarding responsible antibiotic use, animal welfare, environmental sustainability, and the production of safe foods that support human health [1]. The use of antibiotics as growth promoters, together with their inappropriate therapeutic application, has been associated with the emergence and spread of antimicrobial resistance, posing significant threats to both animal and public health [2]. Consequently, considerable efforts have been directed toward identifying natural alternatives that improve productivity and maintain intestinal health without compromising animal welfare or consumer safety. Among these alternatives are prebiotics, probiotics, postbiotics, and bioactive compounds derived from aromatic plants [3]. Maintaining optimal intestinal health and a balanced microbiota is essential for poultry productivity because the gastrointestinal tract regulates several physiological processes, including digestion, immune responses, neuroendocrine signaling, and microbial homeostasis [4].

Functional carbohydrates (FCs) represent a group of nutritional additives with prebiotic properties that differ considerably in their chemical structures and mechanisms of action within the gastrointestinal tract [5–7]. Fructooligosaccharides (FOS) are characterized by β-type linkages (β-2,1 and β-2,6 fructosyl-fructose), which allow their selective fermentation by beneficial bacteria such as Lactobacillus, thereby improving redox status and productive parameters, including feed conversion ratio (FCR) [8–10]. In contrast, mannanoligosaccharides (MOS) and (1,3/1,6) β-glucans, which are derived from the cell wall of Saccharomyces cerevisiae, primarily exert their beneficial effects by inhibiting pathogen colonization and modulating the intestinal microbiome, thereby enhancing antioxidant capacity [1, 11, 12]. More recently, potato resistant starch (RS) has emerged as a structurally distinct FC [6]. RS types II and III possess a semicrystalline α-1,4-D-glucan structure that is resistant to enzymatic hydrolysis [13]. Unlike oligosaccharides, potato RS is preferentially fermented by the intestinal microbiota, promoting the production of butyrate and other short-chain fatty acids (SCFAs), which are essential for maintaining intestinal barrier integrity and gastrointestinal homeostasis [14, 15].

Numerous studies have demonstrated the beneficial effects of individual prebiotic compounds on productive performance, antioxidant status, and intestinal morphology in poultry. Nevertheless, direct comparisons among structurally distinct FCs under identical experimental conditions remain limited. Moreover, the specific differences in microbial modulation induced by MOS, FOS, and potato RS in laying hens have not been clearly elucidated [16]. Although potato RS has attracted considerable attention for its ability to promote SCFA production, particularly butyrate, its effects on intestinal permeability, oxidative status, and cecal microbial ecology in commercial laying hens remain poorly understood. Furthermore, information on how different classes of prebiotic compounds influence the composition and diversity of the cecal microbiota, and how these changes are associated with feed efficiency and intestinal health, remains scarce. Therefore, comparative in vivo evidence is required to better understand the distinct biological responses elicited by these FCs and to establish nutritional strategies to improve gastrointestinal function and productivity in laying hens.

Based on the structural and fermentative differences among these compounds, we hypothesized that MOS, FOS, and potato RS would exert distinct modulatory effects on cecal microbial communities and intestinal physiology. In particular, we expected potato RS to promote a unique microbial profile characterized by enrichment of SCFA-producing taxa, thereby enhancing intestinal barrier function, antioxidant status, and feed efficiency.

Therefore, the present study was designed to comprehensively compare the effects of three structurally distinct FCs, namely MOS, FOS, and potato RS, on productive performance, intestinal health, antioxidant status, and cecal microbial communities in commercial Babcock Brown laying hens during the mid-laying phase. Specifically, the study sought to determine whether the distinct chemical structures and fermentation characteristics of these prebiotic compounds lead to distinct biological responses in the host.

In addition, the study aimed to investigate the effects of dietary supplementation with these FCs on intestinal barrier integrity, intestinal morphology, oxidative status, and goblet cell distribution, as well as to characterize alterations in cecal microbial diversity and taxonomic composition using 16S rRNA gene sequencing. Particular emphasis was placed on identifying diet-associated microbial biomarkers and evaluating whether potato RS preferentially enriches SCFA-producing bacteria associated with improved intestinal homeostasis and feed efficiency. Ultimately, this study aimed to provide comparative in vivo evidence to better understand the relationship between targeted microbial modulation and host physiological responses, thereby supporting the development of effective nutritional strategies for enhancing gastrointestinal health and productivity in commercial laying hens.