Search Results (116)

This study investigated the effects of dietary xylanase and protease supplementation, applied individually or in combination, on growth performance, intestinal characteristics, gut fermentation, meat quality, and skeletal traits in broiler chickens. A total of 540-day-old male broiler chicks were allocated to six experimental groups and fed a control corn-soybean meal-based diet or diets supplemented with xylanase, protease, or a xylanase–protease combination. Enzyme supplementation significantly improved body weight gain and feed efficiency, particularly between days 22 and 42, and reduced intestinal digesta viscosity. Improvements in gut morphology were reflected by increased villus height and villus-to-crypt ratios, accompanied by higher cecal total volatile fatty acid concentrations, increased Lactobacillus populations, and reduced coliform counts. In contrast, breast meat physicochemical composition and antioxidant status were not affected by dietary treatments. Skeletal development was positively influenced, with improvements observed in selected morphometric and structural bone traits. Overall, dietary xylanase and protease supplementation enhanced broiler performance and skeletal development primarily through improved digestive efficiency and favorable modulation of gut morphology, microbial ecology, and intestinal fermentation, without adverse effects on meat quality. Full article

Outcomes of influenza virus infection vary widely across individuals, reflecting not only viral genetics and host factors but also the composition and function of the airway microbiome. Over the past few years, mechanistic work has clarified how specific commensals (for example, Staphylococcus epidermidis and Streptococcus oralis) restrict influenza replication by priming epithelial interferon-λ programs, reshaping intracellular metabolite pools (notably polyamines), dampening host protease activity, and maintaining barrier integrity; meanwhile, pathobionts (notably Staphylococcus aureus and Streptococcus pneumoniae) can enhance viral fitness via secreted proteases and neuraminidases that activate hemagglutinin and remodel sialylated glycoconjugates and mucus, setting the stage for secondary bacterial disease. Recent studies also highlight the gut–lung axis: gut microbiota-derived short-chain fatty acids (SCFAs), especially acetate, protect tight junctions and modulate antiviral immunity in influenza models. Together, these insights motivate translational strategies—from intranasal live biotherapeutics (LBPs) to metabolite sprays and decoy/dual neuraminidase approaches—that complement vaccines and antivirals. We synthesize recent evidence and outline a framework for leveraging the airway microbiome to prevent infection, blunt severity, and reduce transmission. Key priorities include strain-level resolution of commensal effects, timing/dosing windows for metabolites and LBPs, and microbiome-aware clinical pathways for anticipating and averting bacterial coinfection. Overall, the airway microbiome emerges as a tractable lever for influenza control at the site of viral entry, with several candidates moving toward clinical testing. Full article

The intestine is considered a habitat for both bacteria and parasites. In this study, many fecal bacterial isolates and the protozoan Blastocystis sp. were recovered from stool samples of individuals with gastrointestinal conditions. Isolated bacteria were tested for extracellular protease production, and the most potent producer was identified by 16SrDNA gene sequencing as P. aeruginosa FZM498. The enzyme was extracted and purified to electrophoretic homogeneity by the DEAE-Sepharose ion-exchanger and SDS-PAGE revealed a major band at 42.15 KDa. It exhibited maximal activity at 35 °C with thermostability at 60 °C (T_1/2 = 200.04 min). It was most active at pH 8.0 and stable at 5.0–9.5. Enzymatic activity was greatly stimulated in the presence of Fe²⁺ ions, but was repressed by Zn²⁺ and Hg²⁺ ions. Inhibition by PMSF, TLCK, aprotinin, benzamidine, and SBTI protease reagents suggests a serine protease family. The V_max and K_m dynamic constants against azocasein were 36.232 U/mL and 0.0072 mM, respectively. It exhibited the lowest K_m value against the synthetic substrate D-Val-Leu-Lys-pNA among all substrates, indicating a plasmin-like activity. Interestingly, when tested against Blastocystis sp., cysts appeared progressively shrunken, ruptured, and mycelial-like, indicating complete structural collapse with leakage of intracellular contents. The importance of this research is that it is the first study to test the anti-Blastocystis activity of an extracted bacterial serine protease from the gut. This could be a promising, eco-friendly, natural alternative as an anti-Blastocystis agent. The objective of this study was to isolate, purify, and biochemically characterize an extracellular serine protease produced by gut-associated bacteria, as well as to assess its in vitro anti-Blastocystis efficacy as a potential natural and ecologically friendly antiparasitic therapy. Full article

Probiotics hold great potential in aquaculture, as they can effectively modulate gut microbiota and improve fish health, thereby enhancing farming efficiency. Translating this potential into practical application critically relies on screening high-efficacy probiotic strains. This study evaluated the growth-promoting and health-enhancing effects of probiotic candidates Lactobacillus rhamnosus GG (LGG), Lactobacillus plantarum FZU310 (LP-FZU310) and Bacillus subtilis FZU103 (BS-FZU103) in largemouth bass (Micropterus salmoides). After feeding different probiotics for 30 days, the growth, antioxidant, and intestinal enzyme indicators of M. salmoides were detected. BS-FZU103 demonstrated superior efficacy among the tested strains, showing significant differences in both specific growth rate (SGR) (p < 0.05) and condition factor (CF) (p < 0.05). It also markedly enhanced hepatic antioxidant status, elevating superoxide dismutase and glutathione peroxidase activities while reducing malondialdehyde levels by 80%. Improved liver integrity was indicated by significant decreases in serum alanine aminotransferase, aspartate aminotransferase and alkaline phosphatase. Digestively, BS-FZU103 specifically increased intestinal amylase activity by 14.7%, without affecting protease or lipase, suggesting enhanced carbohydrate digestion. 16S rRNA sequencing revealed BS-FZU103 remodeled gut microbiota, increasing Proteobacteria abundance at the phylum level and enriching Bacillus while reducing Clostridium sensu stricto 1 at the genus level. Functional prediction based on PICRUSt2 indicated an enhanced metabolic potential of the gut microbiota, with inferred upregulation of pathways related to carbohydrate transport and metabolism (e.g., ABC transporters) and intestinal enzymatic activities. Collectively, BS-FZU103 is associated with metabolic modulation, promoting M. salmoides growth through gut microbiota remodeling, hepatic antioxidant fortification, and targeted augmentation of carbohydrate utilization efficiency. Full article

This study aims to investigate the differences in microbial community structure between fast-growing (FG) and slow-growing (SG) short-finned eels, Anguilla bicolor pacifica, using high-throughput 16S rDNA sequencing, and to evaluate the potential probiotic properties of Bacillus tropicus isolated from eel intestinal microbiota to enhance growth performance. High-throughput 16S rDNA sequencing revealed no significant differences in the α-diversity between FG and SG eels. Bacterial genera such as Cetobacterium, Clostridium, and Bacteroides were predominant in both groups, with Edwardsiella, Aeromonas, and Fusobacterium being more abundant in SG eels, suggesting a higher presence of potential pathogens. The analysis of the relative abundance of gut microorganisms revealed that SG eels harbored higher levels of potentially pathogenic bacteria, including Edwardsiella tarda and Aeromonas jandaei. In contrast, FG eels exhibited a greater abundance of the potential probiotic B. tropicus. Six strains of bacteria with relative abundance were isolated from the FG group, displaying superior digestive enzyme activity, including protease, lipase, amylase, cellulase, xylanase, and phytase, particularly strain FG2. Phylogenetic analysis confirmed that FG2 was closely related to B. tropicus. A virulence assessment confirmed the non-pathogenic nature of B. tropicus FG2, supporting its probiotic potential. Furthermore, feeding eels a diet supplemented with B. tropicus FG2 significantly enhanced growth performance, as evidenced by increased final weight percentages of weight gain and total production per tank (p < 0.05), while the proximate composition of the dorsal muscle showed an increase in lipid content (p < 0.05). These findings highlight B. tropicus FG2 as a promising probiotic for aquaculture applications. Full article

This study evaluated the effects of matrix-dosed protease supplementation on growth performance, nutrient utilization, intestinal morphology, serum biochemistry, carcass traits, and economic return in broiler chickens. A total of 240 Cobb 500 chicks were assigned to six dietary treatments (T0–T5) with four replicates of 10 birds each for 33 days. Protease supplementation, particularly with protease F at 250 g/tonne (T5), significantly increased body weight at day 7 (163.0 ± 1.4 g; p = 0.002) and day 21 (854.0 ± 7.0 g; p = 0.014), and improved the feed conversion ratio at day 33 (1.54 ± 0.01; p = 0.002). Birds in the T5 group consistently exhibited the highest serum total protein (p < 0.001 on Day 21; p = 0.002 on Day 33), albumin (p < 0.001 on both days), and creatinine (p < 0.001 on Day 21; p = 0.006 on Day 33), along with reduced low-density lipoprotein (LDL) levels (p < 0.001 on Day 21; p = 0.002 on Day 33). Intestinal morphology was also enhanced, with villus height increasing to 874.0 ± 1.0 µm at day 21 and 931.0 ± 1.0 µm at day 33, accompanied by greater villus height-to-crypt depth ratios (11.23 ± 0.02 and 12.59 ± 0.01, respectively; p < 0.001). Moreover, apparent ileal digestibility of dry matter, crude protein, metabolizable energy, and amino acids were improved in T5 compared with the control and other treatments. Economic analysis showed the highest profit and return on investment (7.01%) in T5, followed by T4 and T2. These findings indicate that matrix-based protease supplementation enhances growth, nutrient absorption, and gut morphology while delivering substantial economic benefits, making it a cost-effective strategy for improving broiler productivity and profitability in commercial production systems. Full article

Objectives: This experiment investigated the response of carcass composition, digestive function, hepatic lipid metabolism, intestinal microbiota, and serum metabolomics to excessive or restrictive dietary energy in Ningxiang pigs. Methods: A total of 36 Ningxiang pigs (210 ± 2 d, 43.26 ± 3.21 kg) were randomly assigned to three treatments (6 pens of 2 piglets each) and fed a control diet (CON, digestive energy (DE) 13.02 MJ/kg,), excessive energy diet (EE, 15.22 MJ/kg), and restrictive energy diet (RE, DE 10.84 MJ/kg), respectively. Results: Results showed that EE significantly increased the apparent digestibility of crude protein and total energy (p < 0.01), as well as the activities of jejunum neutral protease and ileal lipase (p < 0.05). With the increase in energy level, the apparent digestibility of ash, dry matter, and ether extract significantly increased (p < 0.01). RE significantly increased high-density lipoprotein cholesterol (HDL-C) content, significantly decreased triglycerides (TG), free fatty acid (NEFA), and total cholesterol (TC) contents, and up-regulated lipoprotein lipase (LPL) mRNA expression in the liver (p < 0.05). EE significantly increased the hepatosomatic index, the contents of low-density lipoprotein cholesterol (LDL-C) and total bile acids (TBA), and significantly up-regulated the mRNA expression of lipogenic genes acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), and sterol regulatory element-binding protein-1C (SREBP-1C) in the liver (p < 0.05). The abundance of p_Firmicutes was significantly increased and the abundance of p_Bacteroidetes was significantly decreased in test groups, while the ratio of the two was significantly increased in the RE group (p < 0.05). EE also significantly increased the abundance of g_Clostridium_sensu_stricto_1 (p < 0.05). The identical serum differential metabolites between the EE and RE group belong to phosphatidylcholine (PC), mostly being up-regulated in the EE group and down-regulated in the RE group (p < 0.05), one of which was mapped to the pathway of glycerophospholipid metabolism (KEGG ID: C00157). The relative content of serum trimethylamine N-oxide (TMAO, a microbial metabolite) was significantly decreased in the EE group (p < 0.05). Conclusions: The findings suggest RE had no obvious negative effect on carcass traits of Ningxiang pigs. Apart from exacerbated body fat deposition, EE promoted fat accumulation in the liver by up-regulating the expression of lipogenic genes. Dietary energy changes affect hepatic bile acid metabolism, which may be mediated through the glycerophospholipid metabolism pathway, as well as disturbances in the gut microbiota. Full article

Insect establishment and dispersal are often influenced by temperature, with gut microbiota playing a critical role in host adaptation to environmental stress. This study investigated how gut bacterial structure and function in the invasive red-haired bark beetle (RHB), Hylurgus ligniperda (Fabricius) respond to temperature fluctuations, focusing on three core culturable bacteria: Rahnella perminowiae, Serratia marcescens, and Hafnia psychrotolerans. We found that temperature variations induced specific structural changes in the gut bacterial community, which in turn affected key functional processes such as carbohydrate metabolism. Notably, the relative abundance of Rahnella increased by more than 10% during the cold period (CP), and it maintained stable production of proteases and lipases under low temperatures—a trait that may be crucial for supporting host development in cold environments. Feeding on the diet converted by R. perminowiae at 5 °C resulted in a 20.9-day reduction in pupation time and a 1.8-fold increase in adult body mass compared to the blank control group, respectively. We propose that temperature remodels the gut microbiota by modulating competitive relationships among functional bacteria. This regulatory mechanism, based on functional redundancy and dynamic balance, serves as a buffer strategy that aids insect adaptation to temperature changes. Our findings provide new insights and a theoretical foundation for understanding pest outbreak patterns under climate warming and developing microbe-targeted control strategies. Full article

Metarhizium anisopliae, an entomopathogenic fungus, can produce four extracellular proteases, subtilisin (Pr1), trypsin (Pr2), metalloproteases (Pr3), and cysteine proteases (Pr4), which are important for pathogenicity of M. anisopliae in target hosts. In order to understand their function in M. anisopliae pathogenicity, third-instar nymphs of Locusta migratoria were fed with a diet containing either conidia of M. anisopliae strain IPPM202 or in combination with one of the four inhibitors (TPCK: tosyl-phenylalanine chloromethyl-ketone, inhibitor of Pr1; EDTA: ethylenediaminetetraacetic acid, inhibitor of Pr3; APMSF: 4-amidinophenyl methanesulfonyl fluoride, inhibitor of Pr2; CI1: cathepsin inhibitor 1, inhibitor of Pr4). The effects on mortality, midgut integrity, and the gut enzymes peroxidase (POD), catalase (CAT), superoxide dismutase (SOD), and phenol oxidase (PO) were examined. The results indicated that exposure to IPPM202/TPCK and IPPM202/CI1 caused decreased mortality to L. migratoria with no loss of midgut epithelial cellular integrity. On the other hand, exposure to IPPM202/APMSF or IPPM202/EDTA mixtures resulted in higher mortality similar to PPM202, with severely damaged epithelial gut cells with fragmented microvilli, broken endoplasmic reticulum, and disrupted nucleus membrane. The activity of the protective enzymes POD, SOD, CAT, and PO all increased significantly when L. migratoria was treated with IPPM202 only, but decreased when any one of the inhibitors was added. We further concluded that TPCK, a subtilisin (Pr1) inhibitor, and CI1, a cysteine protease (Pr4) inhibitor, played important roles in the pathogenicity of the M. anisopliae strain IPPM202. Conversely, trypsin (Pr2) and metalloproteases (Pr3) did not have a role in the given process. We further concluded that trypsin (Pr2) and metalloproteases (Pr3) do not contribute to the fungal infection process, while the subtilisin (Pr1) inhibitor TPCK and cysteine protease (Pr4) inhibitor CI1 play critical roles in the pathogenicity of Metarhizium anisopliae strain IPPM202, thus providing a foundation for targeted biocontrol strategies. Full article

Background/Objectives: Global warming and concomitant extreme weather events have markedly increased the incidence of heat stroke. Heat stroke (HS) poses a substantial threat to cerebral health by triggering neuroinflammation and accelerating neurodegenerative processes. The activation of the Nod-like receptor protein 3 (NLRP3) inflammasome for interleukin-1β (IL-1β) secretion has been implicated as a critical mechanism underlying HS-related fatalities. However, the potential role of specific dietary factors to counteract heat stroke-induced neurotoxicity remains largely underexplored. We previously reported that eicosapentaenoic acid (EPA) and urolithin A (UroA), a gut metabolite of ellagic acid, effectively suppress NLRP3 inflammasome activation against metabolic or pathogenic insults. This study aimed to assess the impact of eicosapentaenoic acid (EPA), urolithin A (UroA), and their combination on mitigating heatstroke-mediated NLRP3 inflammasome activation in microglial cells. Methods: In vitro heatstroke conditions were replicated by subjecting murine BV2 microglial cells to a high temperature (41 °C) under hypoxic conditions. To achieve nutrient loading, BV2 cells were preincubated with either EPA (50 µM) or UroA (10 µM). NLRP3 inflammasome activation was evaluated by proinflammatory gene expression, caspase-1 cleavage in cells, and IL-1β secretion to the medium. The caspase-1 activation was determined using a luciferase-based inflammasome and protease activity reporter (iGLuc) assay. Results: Exposure to high temperatures under hypoxia successfully mimicked HS conditions and promoted NLRP3 inflammasome activation in BV2 cells. Both EPA and UroA substantially attenuated the heat stroke-induced priming of proinflammatory genes. More importantly, EPA and UroA demonstrated a synergistic effect in mitigating HS-induced active caspase-1 production, leading to a dramatic decrease in IL-1β secretion. This synergistic effect between EPA and UroA was further confirmed by the iGLuc reporter assay. Conclusions: Dietary enrichment with EPA and UroA precursors may constitute an efficacious strategy for mitigating heat stroke-mediated neuroinflammation and neurodegenerative diseases. Full article

The aim of this study was to conduct a comparative analysis of the effects of native quinoa grain with a high saponin content and quinoa grain subjected to preliminary saponin removal with low saponin content on growth, meat quality, biochemical blood composition, and the expression of genes related to muscle growth, gut health, and nutrient transport in broiler chickens. The control group of chickens received a standard diet. The SAP group feed contained quinoa grain without saponin removal (saponin level—5.20%) at 3% of the “Starter” feed mass and 5% of the “Grower” and “Finisher” feeds, maintaining the same nutritional values as the control group. The SAP-FREE group feed contained quinoa grain that was pre-treated to remove saponins by washing with water for 60 min at a temperature of 50 °C (saponin level—0.24%) in the same amount as the SAP group. The research results indicated certain advantages of unprocessed quinoa grain in relation to saponin content. Specifically, in the SAP group, the broiler performance index was at the same level as the control, while the SAP-FREE group had a high mortality rate (10%), resulting in a performance index that was 23.82 units lower than the control. The use of quinoa grain with high saponin content promoted better development of thigh muscles by 9.6% compared to the control (p = 0.008) and increased yields of wing, neck, and back muscles by 2.9 abs.% (p = 0.007) compared to the use of purified quinoa grain. The fat yield decreased by 1.7 abs.% (p = 0.015) with saponin-free quinoa compared to the control and by 2% (p = 0.008) compared to the high saponin group, making this feeding system viable for producing dietary meat. Upon stopping the feeding of purified quinoa, chickens showed a 34.0% increase in AST activity (p = 0.019) and a 15.7% increase in creatinine levels (p = 0.008), likely indicating intensified protein metabolism upon cessation of the inhibiting factor of purified quinoa. Molecular genetic studies revealed a 1.6-fold increase in IGF1 gene expression (p = 0.014) in breast muscle and a 69.12-fold increase (p = 0.010) in AvBD9 in the cecum due to high-saponin quinoa grain, while purified quinoa increased GHR gene expression by 3.29 times (p = 0.039) in breast muscle and decreased IRF7 activity to 2^−ΔΔCT = 0.54 (p = 0.017). The expression of transporter protein genes decreased to low or undetectable levels, indicating the presence of anti-nutritional factors and the need for further research on feeding quinoa with the addition of proteases. Thus, high-saponin quinoa grain, unlike purified quinoa, positively influences gut health and bird survival, maintaining performance levels similar to the control, suggesting the feasibility of using unprocessed quinoa in poultry nutrition, thus avoiding additional costs in feed preparation. Full article

The analysis of proteins in stool samples can significantly enhance the study of mammalian physiology and disease. In this study, we investigated the fecal proteome of clinically healthy dogs (n = 26) by a label-free proteomics approach to evaluate the impact of breed differences. The dogs were divided into two groups (n = 13 each) based on their breed, specifically Weimaraner and Dalmatian, the former known for their possible susceptibility to gastrointestinal disease. Quantitative and qualitative differences between the two experimental groups were identified based on analyses performed on pooled biological samples. The overall fecal proteome profile comprised 58 proteins, of which 37 were common, while comparative proteomics analysis detected 15 proteins with different abundances. Notably, the fecal proteome of Weimaraners showed an over-representation of proteins such as pantetheinase, which promotes inflammatory reactions; ferritin heavy chain and hemoglobin, possibly associated with gut ulceration and/or rectal bleeding typical of IBD; and anionic trypsin, implicated in inflammatory bowel disease. Notably, in Dalmatians, despite the absence of specific predispositions, some proteins associated with chronic enteropathy (e.g., carboxypeptidase B and serine protease 1) were also over-represented. Additionally, some proteins linked to breed variation included enzymes associated with “protein digestion and absorption” and “glycolysis and gluconeogenesis”. These findings suggest, for the first time, that the variable breed is a factor that may potentially influence the fecal proteome in dogs. Full article

COVID affects around 400 million individuals today with a strong economic impact on the global economy. The list of long COVID symptoms is extremely broad because it is derived from neurological, cardiovascular, respiratory, immune, and renal dysfunctions and damages. We review here these pathophysiological manifestations and the predictors of this multi-organ pathology like the persistence of the virus, altered endothelial function, unrepaired tissue damage, immune dysregulation, and gut dysbiosis. We also discuss the similarities between long COVID and vaccine side effects together with possible common immuno-inflammatory pathways. Since the spike protein is present in SARS-CoV-2 (and its variants) but also produced by the COVID vaccines, its toxicity may also apply to all mRNA or adenoviral DNA vaccines as they are based on the production of a very similar spike protein to the virus. After COVID infection or vaccination, the spike protein can last for months in the body and may interact with ACE2 receptors and mannan-binding lectin (MBL)/mannan-binding lectin serine protease 2 (MASP-2), which are present almost everywhere in the organism. As a result, the spike protein may be able to trigger inflammation in a lot of organs and systems similar to COVID infection. We suggest that three immuno-inflammatory pathways are particularly key and responsible for long COVID and COVID vaccine side effects, as it has been shown for COVID, which may explain in large part their strong similarities: the renin–angiotensin–aldosterone system (RAAS), the kininogen–kinin–kallikrein system (KKS), and the lectin complement pathway. We propose that therapeutic studies should focus on these pathways to propose better cures for both long COVID as well as for COVID vaccine side effects. Full article

Two trials evaluated the effects of dietary protease inclusion in weaned piglets fed diets with or without crude protein (CP) reduction, focusing on performance, intestinal health, and amino acid digestibility. In Trial I, 270 piglets (21–63 days) received six treatments: control (PC), PC with 100 g/ton protease A (PC+A), CP reduced by 1.0% (NC1) or 1.5% (NC1.5), NC1.5 with 50 g/ton protease A (NC1.5+A), and NC1.5 with 50 g/ton protease B (NC1.5+B). PC+A improved weight gain, feed intake, and feed conversion compared with NC1.5+A. The incidence of diarrhea was reduced in animals fed protease-supplemented diets (PC+A, NC1.5+A and NC1.5+B). PC had greater ileal villus height than NC1.5+B, and PC+A showed a higher jejunal villus-to-crypt ratio than reduced CP groups. NC1.5+B increased jejunal expression of IL-6, TNF-α, and haptoglobin. In Trial II, 12 ileal-cannulated piglets received diets with or without protease A. Protease improved the standardized ileal digestibility (SID) of methionine+cysteine and tryptophan but reduced the SID of glycine and proline. While protease supplementation can improve some amino acids (Met+Cys and Thr) protein digestibility, our findings suggest it cannot fully replace careful amino acid balancing in CP-reduced diets. However, protease-supplemented diets were associated with improved intestinal morphometry and a reduced incidence of diarrhea. Full article

The objective of this study was to evaluate the effects of various feed additives on odor emissions, gut health, and stress responses in laying hens fed low-protein diets. Four commercially available functional feed additives (Bacillus subtilis, protease, saponin, and thyme-based essential oil) were selected for this study. A total of 288 Hy-Line brown laying hens aged 49 weeks were randomly fed on one of six experiment diets: a 16% standard crude protein diet, a 12% low-crude-protein (LCP) diet, and LCP diets supplemented with Bacillus-based probiotic, protease, saponin, or thyme-based essential oils prepared for 8 weeks. Each treatment had eight replicates with six birds per replicate. Lowering crude protein levels affected the laying performance, nitrogen balance, odor production (i.e., ammonia), and nutrient digestibility but did not alter eggshell quality or fecal short-chain fatty acids. Dietary additives added into the LCP diet did not affect the laying performance, egg qualities, and nitrogen balance but increased crude ash digestibility compared with the LCP-diet-fed laying hens. Branched-chain fatty acids tended to be higher in all laying hens fed low-CP diets, irrespective of feed additives. Notably, low vs. standard protein diets tended to increase yolk corticosterone levels, which is an indicator of stress responses in chickens. This low-CP-mediated increase in yolk corticosterone was partially decreased by 20.8–48.6% on average, depending on the additives used. Our study suggests that low-protein diets could effectively lower nitrogen excretion and odor emissions. However, adding dietary additives into low-protein diets has minimal effects on low-CP-diet-fed laying hens, which needs further studies to clarify the role of low-crude-protein diets and dietary additives in modulating hindgut fermentation via shaping the gut microbiota and stress responses of laying hens. Full article