Animal welfare, ecological sustainability, and controlled usage of medical pharmaceuticals are all objectives for producing healthy products devoid of chemical residues or pharmaceutical drugs (1). Recently, along with implementing antibiotics bans in many countries, several studies have been conducted to search for alternatives to antibiotics in the livestock industry (2) due to increased interest in human nutrition (3).

Since ancient times, honey bee venom (BV) has been frequently used as a painkiller and treatment for inflammatory diseases Yoon et al. (4). Son et al. (5) reported that more than 40 pharmaceutically active components are considered in bee venom, including peptides and enzymes, many of which have yet to be investigated (1). In several investigations, bee venom has been established to have anti-inflammatory and antibacterial characteristics. Pucca et al. (6) describe bee venom as comprising low molecular polypeptides and a complicated mixture of amino acids, peptides, proteins, enzymes, sugars, biogenic amines, volatile acids, phospholipids, and pheromones. In addition, the BV contains biologically active amines, such as histamine, adrenaline, and non-peptide derivatives, with a range of pharmacological properties. Melittin, apamin, and adolapin (polypeptides) act as anti-inflammatory and antibacterial factors and are the primary elements of BV (7). Melittin is the primary active component that enhances the pituitary and adrenal glands to produce catecholamine and cortisone, that has anti-inflammatory properties (8). Bee venom can be used safely and healthily in the rabbit industry instead of drugs (9). Melittin inclusion in the rabbits’ diets can increase their production efficiency (10) and could also be used for the prevention of disease (Apitherapy) (11). BV treatment provides a novel option for antimicrobial growth promoters in the nutrition of animals (11). On the other hand, the anti-oxidant, antibacterial, and analgesic properties of BV were also discovered by Zhang et al. (12). Moreover, because apamin and phospholipase A₂ in BV have potent immuno-regulatory activity, several studies have demonstrated that BV therapy could help treat multiple immune disorders (13).

As a consequence of improper use, tetracycline residues may be detected in marketable animal products, representing a risk to humans and causing allergic reactions. Furthermore, prolonged use of low-dose antibiotics in products can contribute to the proliferation of drug-resistant microbes (14). As a result, BV significantly affects broiler performance and could be a feasible alternative to antimicrobial growth promoters to prevent the use of antimicrobials regularly (11).

For more than 60 years, antibiotics have been allowed to be used as growth promoters in poultry feeds without a prescription (15). Recently, many studies have been conducted to search for an alternative to antibiotics in the livestock industry. This ban is due to the fact that antibiotics as growth promoters in feed additives have led to the formation of resistant bacterial strains and antibiotic residues in animal products (16). From this perspective, medicinal plants can be used as an alternative to antibiotics and improve poultry live performance, such as extracted essential oils (17).

The modern trend in poultry feeding is to search for alternatives to antibiotics as growth promoters in poultry rations due to the harmful effects of antibiotic residues in poultry meat on human health. Most of the recent research tended to study bee venom to improve growth in broiler chicks, and there is a significant lack of studying the effect of bee venom on growing rabbits. This research investigated the impact of replacing BV instead of Oxytetracycline in growing rabbits’ drinking water on the productive performance, biochemical blood constituents, anti-oxidant status, immune response, and cecal microbial count of growing rabbits.

In the current investigation, rabbits were handled under the recommendations of the Pharmaceutical and Fermentation Industries Development Center, City of Scientific Research and Technology Applications (SRTA-City), Alexandria, Egypt, with the approval of the Institutional Animal Care and Use Committees (IACUCs)/IACUC # 37-6F-1021.

Regarding the BV collection method, it can be summarized that the electric method is more effective than the manual method, as the active ingredients were present in a higher concentration in BV from the electric technique than the manual method. In our study, the primary substances in BV were melittin, protease, hyaluronidase, and phospholipase A2; this result was in accordance with Teoh et al. (26) and Mammadova and Topchiyeva (27). According to Damianoglou et al. (28), approximately 60 components have been identified in bee venom, with melittin being the most abundant. In the same context, several studies mentioned that Honey bee venom comprises many compounds, including enzymes, proteins, peptides, and numerous other tiny substances (amino acids, catecholamines, sugars, and minerals) (29). According to El-Hanoun et al. (9), the active substance in BV may be a significant external environmental component that can directly impact animal welfare.

Increased daily BWG and decreased daily FI of BV rabbits’ groups reflected in FCR that was significantly improved (p ≤ 0.01) versus the control and OXT groups. A previous study by Han et al. (11) demonstrated that at 28 days of age, adding BV to broiler drinking water significantly increased BWG compared to the control group (p < 0.05); as well, FI was numerically higher but without significance. So, BV can potentially replace antimicrobial growth promoters in broiler nutrition. This is in agreement with Han et al. (30), who indicated BV injection resulted in a net increase in BW gain and survivability in piglets. According to Rabie et al. (10) and El-Hanoun et al. (9), BV substance can stimulate productive performance, which could be ascribed to its health benefits, including the prevention/treatment of diseases (Apitherapy) (8). Additionally, it has antimicrobial properties (11). Likewise, stinging or subcutaneously injecting piglets with BV increased LBW significantly with decreased FI. Moreover, BV improved the survivability of the piglets, so it was possible to use BV as an alternative to antibiotics growth promoters (30, 31). In contrast, Ahmad et al. (32) stated that the FI of birds supplemented with BV was significantly higher than that of control birds. Still, this increase was positively correlated with the increasing BWG of the treated birds. On the other hand, injecting broilers with BV extract did not affect LBW at 42 days and BWG from 0 to 6 weeks; additionally, FI was not significantly lower in the treated groups than in the control group (33). Moreover, according to Lu et al. (34), melittin does not harm the immune system, which should be another contributing factor to growth performance in a highly concentrated production environment in the broiler industry. It seems likely that BV strengthens the immune response to the normal environmental, social, and nutritional challenges faced by newly born chicks.

Carcass characteristics (Hot carcass and Total edible parts) improvement that was noticed in the BV treated groups was attributed to the improvement that happened in the growth performance of BV rabbits compared to the control group, whereas BWG was significantly increased and FI significantly decreased, which in return was reflected on FCR improvement, which referred to BV boosted rabbit ability to convert feed to meat. On the other hand, the data presented in Table 4 illustrated an increase in the digestibility Coefficient and Nutritive Values for BV groups in comparison to the control group. These results indicated increased nutritive absorption values through the intestine; consequently, there was an abundance of nutrients to build muscles. This hypothesis is consistent with the increased small intestine weight in the BV groups in contrast to the control groups. In the same context, the increased pancreas weight of BV rabbits more than the control group indicates an increase in pancreatic function of insulin hormone secretion, which plays a vital role in carbohydrate metabolism.

Moreover, increased abdominal fat relative weight and decreased blood Trgly concentration of BV treated groups relative to the control group can be explained by the fact that BV affected lipid metabolism and stimulated and enhanced the lipid transfer process from blood to be stored in the adipose tissue. Our results agree with that found by El-Kholy (35), who reported that the significant improvement in carcass characteristics for treated groups could be due to the treated rabbits’ improved growth performance, which mirrors the rabbits’ health. Furthermore, Lu et al. (34) mentioned that melittin contributed to growth performance in intensive broiler production systems. Besides, the BV boosted an immune response to encounter the challenges of the newly born chicks’ normal environmental and nutritional conditions. Also, BV supplementation counterbalanced stress within days after birth. In addition, no morphological injury signs to the studied organs were observed under BV treatment, which was consistent with that found in broilers treated with BV in drinking water (11).

Increasing plasma TP in the present study agreed with that obtained by El-Hanoun et al. (9) and Elkomy et al. (36), who demonstrated that injected adult male and female rabbits with BV significantly increased blood TP and this effect was dependent on BV dosage (37). BV induced the immune system to increase plasma Total Glob concentration and immune-globulin fractions (IgG and IgM), which improved rabbit health. Our results were consistent with those of Elkomy et al. (36), El-Hanoun et al. (9), and Gurafi (37), who stated that the BV treatment increased plasma Glob levels significantly more than the control group. This increase may be attributable to the activation of the immune system due to the antigenic properties of BV’s active peptide content (38). In contrast, Ali and Mohanny (33) showed that injecting broiler chicks with BV extract had an insignificant impact on total protein, albumin, and globulin. Likewise, Han et al. (11) mentioned that significant impacts on total protein, albumin, and globulin were noticed due to adding BV via drinking water to chicks. The reduction in AST and ALT enzymes activities that happened in our study following the findings of El-Hanoun et al. (9) and Elkomy et al. (36), who had a similar result when injected subcutaneously with male and female adult rabbits with BV, and this effect revealed an improvement in liver function. Opposite results were found by Han et al. (11); no significant impacts on AST and ALT were observed in broilers receiving BV in their drinking water. In addition, Bolarinwa et al. (1) demonstrated that stinging with BV had no effect on AST and ALT levels when comparing the control and investigated treatments. These previous findings suggest that the impact of BV on the AST and ALT enzyme activities may be dose-dependent, and a high BV dose is needed to decrease the activity of liver enzymes.

Our results revealed that TChol and the harmful part of Chol (LDL and VLDL Chol) were significantly decreased, and the beneficial Chol (HDL Chol) was increased, as well, plasma Trgly level was decreased significantly and this decrease was associated with increased abdominal fat deposition. Decreasing TChol and Trgly levels described in the current study are consistent with El-Hanoun et al. (9), who mentioned that injecting rabbit’s male subcutaneously with BV gradually and significantly decreased plasma Total lipids and TChol. In the same context, BV decreased plasma TChol and Trgly content in mice (39). The same result was found by Mousavi et al. (40) in diabetic rats and revealed that BV significantly reduced the elevated levels of Trgly and TChol to normal levels. In contrast, Han et al. (11) illustrated that BV supplementation via drinking water had no significant impacts on TChol. The relationship between abdominal fat and plasma lipids could be explained by Whitehead and Griffin (41), they found that body fat content correlated closely with plasma VLDL in addition to LDL triglycerides, and it could be used this relationship for selecting leanness in broiler breeding programmers on the basis of measurements of plasma VLDL plus LDL triglyceride. Wayne et al. (42) reported that abdominal fat and blood plasma triglycerides were significantly correlated with body fat. These correlation coefficients indicated that a different relationship exists between fatness and plasma triglyceride concentration in male and female turkeys.

The present results of the effect of BV on redox status agree with that of Elkomy et al. (36), who studied the impact of different BV doses on TAC, GSH, GST, GPx, and SOD. He also found a gradual and significant rise in TAC and anti-oxidative enzyme activities versus the control group, while MDA and TBARS were significantly decreased. Increased anti-oxidant levels due to BV treatment increased the examined does’ resistance to biological and environmental stresses. In the same context, the findings of El-Hanoun et al. (9) on male rabbits, Kim et al. (43), and Han et al. (11) on broiler chickens revealed that BV increased the anti-oxidant profile. According to the findings of Crott and Fenech (44) and Kim et al. (43), BV possesses pharmacological anti-oxidant properties and can protect against cellular injury, thereby reducing oxidative damage to lipids, proteins, and DNA. Kocyigit et al. (45) found a significant increase in total anti-oxidant status (TAS), total oxidant status (TOS), and oxidative stress index (OSI) in rats treated with BV. From previous studies, the SOD, CAT, and GPx are essential enzymes that play an endogenous anti-oxidant protection system and play a crucial part in removing free radicals and maintaining the intracellular redox balance (46). El-Speiy et al. (47) stated that the rise in GPx and SOD activity was indicative of improving anti-oxidant capacity. Furthermore, blood MDA concentration was used as an indicator of lipid peroxidation in the body (48); consequently, the stage of cell injury and lipid peroxidation can be determined by measuring the concentration of MDA, which are the principal products of polyunsaturated lipid peroxidation (46). Based on the previous mentions, the improved TAC, GPx, CAT, and SOD components in the BV groups reflect the fact that BV has anti-oxidant properties that enhance the anti-oxidant defense against oxidative damage, resulting in a decrease in lipid peroxidation markers such as MDA concentration.

Data revealed that BV boosted IgM and IgG levels, and the low-dose BV had the main effect on alerting the immune system through the impact of BV on immuno-globulin fractions, which decreased with the increase of BV dosage. Our findings align with prior research by Elkomy et al. (36) in rabbit does and El-Hanoun et al. (9) in rabbit bucks; they reported that injecting BV led to elevate IgG, IgA, and IgG levels significantly. According to Ali and Mohanny (33), BV as a natural product could stimulate and improve broiler chickens’ immune response without adversely affecting productive performance, possibly because of superoxide production (11). Depending on its pharmacological components, BV can boost the immune system of animals, including cortisone production and antibody creation, and influence cytokine production (5, 49, 50). Bee venom is a blend of biochemicals ranging from small peptides and enzymes to biogenic amines. It is capable of triggering severe immunologic reactions owing to its allergenic fraction. Venom components are presented to the T cells by antigen-presenting cells within the skin. These Th2 type T cells then release IL-4 and IL-13, which subsequently direct B cells to class switch to the production of IgE. BV can induce elevation of specific IgE and IgG antibodies (51) and leads to the production of IgE antibodies, which can respond to various antigens (52).

Data on cecum microbial count and differentiation agreed with the previously detected results. Previous studies have mentioned that BV is a mixture of substances and has bioactive properties, including growth promotor (31), anti-inflammatory (53), and antimicrobial (11) in treated animals with no side effects detected on them. According to Leandro et al. (54), the BV antimicrobial activity may be due to several peptides, which involve melittin, apamin, adolapin, enzymes, biologically-active amines, and non-peptide parts. Above that, BV contains Phospholipase A2 (PLA2), which has antibacterial properties, plus mellitin contributes to this antibacterial effect (55, 56). The same result was reported by Dani et al. (57) and Permual et al. (58), who noted that Mellitin is a primary component of BV and has antibacterial properties. The data obtained by Zolfagharian et al. (59) showed that BV inhibited bacterial growth and had stronger antibacterial efficacy than the gentamicin antibiotic. Consequently, BV has antimicrobial properties against gram-positive as well as gram-negative bacteria, including Escherichia coli (E. coli), Salmonella spp., Enterobacter cloacae, Staphylococcus aureus (S. aureus), and coagulase-negative Staphylococcus (60). Moreover, numerous impacts on distinct cell types involving antiviral and anti-inflammatory effects have been observed (59). Ortel and Markwardt (61) also mentioned that microorganisms were also found to be more sensitive to BV at lesser concentrations.

In conclusion, bee venom (BV) at 2, 4, and 8 mg dosages led to a significant increase in LBW and BWG accompanied by a decrease in FI, resulting in improved FCR and carcass traits. Moreover, BV positively affects the immunological and anti-oxidative responses and reduces the pathogenic bacteria count in the hindgut in growing rabbits.

Bee venom can be an effective, safe, and healthy alternative to antibiotics for use in rabbit farms to improve the productivity and health status of growing rabbits and reduce the activity of pathogenic bacteria in the cecum.

The original contributions presented in the study are included in the article/supplementary material. Further inquiries can be directed to the corresponding authors.

The animal studies were approved by Animals were handled in the present study following the guidelines of the Pharmaceutical and Fermentation Industries Development Center, City of Scientific Research and Technology Applications (SRTA-City), Alexandria, Egypt, after the approval of the Institutional Animal Care and Use Committees (IACUCs)/IACUC # 37-6F-1021. The studies were conducted in accordance with the local legislation and institutional requirements. Written informed consent was obtained from the owners for the participation of their animals in this study.

AE, ME-S, SH, and TS: supervision, project administration, conceptualization, methodology, formal analysis, supervision, and writing–original draft. ME-B: resources. AE, HaE, and TS: conceptualization, methodology, formal analysis, and investigation. MW, SC, IK, and HoE: supervision, visualization, data curation, and writing–review and editing. All authors contributed to the article and approved the submitted version.

This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-RS-2023-00275307).