Search Results (285)

This study evaluated the effects of adding a Saccharomyces cerevisiae fermentation product (SCFP) to adult cat diets on palatability, intestinal health, nutrient digestibility, immune parameters, and the fecal microbiome over 42 days. Sixty-three healthy Domestic Short-hair cats were randomized to three diets: a control diet (CD) without SCFP, or the same diet containing 1.0% or 2.0% SCFP, targeting daily intakes of 150 or 300 mg/kg body weight, respectively. Body weight and blood parameters remained within reference ranges across all groups. Stool quality was largely not affected, although compared with controls, SCFP-supplemented cats had slightly firmer stools at day 21, and increased fecal pH. Shotgun metagenomic sequencing revealed that microbiome diversity remained steady in SCFP-fed cats, whereas diversity in the control group declined over time. Fecal immunoglobulin A concentrations were lower in cats supplemented with SCFP at 150 mg/kg BW by the end of the study, and positive shifts in the circulatory leukocyte profile were observed at both inclusion levels. Apparent total tract macronutrient digestibility did not differ among groups. Palatability tests showed diets with SCFP were generally preferred, indicating a potential benefit for commercial feline feeds, particularly at the 150 mg/kg BW level, which was preferred over 300 mg/kg BW. Overall, these findings suggest that SCFP can act as a functional ingredient in feline nutrition to maintain microbial diversity and enhance diet acceptance without compromising digestibility. Full article

This study aimed to investigate the effects of dietary selenium (Se) supplementation levels on the apparent total tract digestibility (ATTD) of crude protein, crude fat, and crude ash, and cecal microbiota diversity and composition in nursery piglets challenged with deoxynivalenol (DON). Twenty-four weaned piglets were allocated to four dietary treatment groups (n = 6). The four treatment diets were as follows: (1) post-weaning diet containing 0.3 mg/kg of Se; (2) post-weaning diet without supplemental Se + 3 mg/kg DON; (3) post-weaning diet containing 0.3 mg/kg Se + 3 mg/kg DON; and (4) post-weaning diet containing 0.5 mg/kg Se + 3 mg/kg DON. The native Se levels in the feedstuffs were 0.03–0.06 mg/kg. The Se source was supplemented in the form of selenomethionine. The trial lasted 28 days, with fecal samples collected during the final 4 days for ATTD analysis of crude protein, crude fat, and crude ash. Upon trial completion, cecal digesta was sampled for microbial diversity and composition analysis. The results showed that piglets in the 0.5 mg/kg Se + DON group had a higher ATTD of crude fat than the 0 mg/kg Se + DON group and a greater ATTD of crude ash than the 0.3 mg/kg Se + DON group (p < 0.05). Additionally, the 0.5 mg/kg Se + DON group displayed increased Shannon and Chao 1 indices compared to the 0 mg/kg Se + DON group (p < 0.05). Microbial composition analysis revealed higher Firmicutes abundance in the 0.5 mg/kg Se + DON group than in the 0.3 mg/kg Se + DON group, along with reduced Euryarchaeota abundance relative to the 0 mg/kg Se + DON group (p < 0.05). The 0.5 mg/kg Se + DON group also had higher Selenomonadaceae abundance than the other groups (p < 0.05). Pearson correlation analysis indicated a negative correlation between Selenomonadaceae abundance and the ATTD of ash, whereas [Eubacterium]_coprostanoligenes_group abundance was positively correlated with the ATTD of ash (p < 0.10). Collectively, dietary Se supplementation at 0.5 mg/kg improved nutrient digestibility in DON-exposed nursery piglets, which was potentially associated with the modulation of cecal microbiota diversity and composition. Full article

The use of exogenous enzymes in the nutrition of dairy cows is an innovative and efficient strategy to maximize productivity and milk quality, with positive applications in the economic and environmental aspects of dairy farming. Therefore, the objective of this study was to evaluate whether the addition of a blend of exogenous enzymes to the diet of lactating Jersey cows has a positive effect on productive performance, milk quality, animal health, ruminal environment, and digestibility. Twenty-one primiparous Jersey cows, with 210 days in lactation (DL), were used. The exogenous enzymes used were blends containing mainly protease, in addition to cellulase, xylanase, and beta-glucanase. The animals were divided into three groups with seven replicates per group (each animal being the experimental unit), as follows: Control (T-0), basal diet without enzyme addition; Treatment (T-80), animals fed enzymes in the diet at a daily dose of 80 mg per kg of dry matter (DM); Treatment (T-160), animals fed enzymes in the diet at a daily dose of 160 mg per kg of DM. The study lasted 84 days, during which higher milk production was observed in the treated groups (T-80 and T-160) compared to the control group (p = 0.04). When calculating feed efficiency from days 1 to 84, greater efficiency was observed in both groups that received the blend compared to the control (p = 0.05). In the centesimal composition of the milk, it was observed that the percentage of protein in the milk of the T-160 group was higher compared to the control group (p = 0.03). The effect of the enzymes was verified for butyric (p = 0.05) and palmitic (p = 0.05) fatty acids. We also observed the effect of the enzyme blend on the amount of volatile fatty acids (VFAs), which were higher in the ruminal fluid of cows that received the enzymes (p = 0.01). Cows that consumed enzymes showed a higher apparent digestibility coefficient of crude protein (p = 0.01). In vitro, the main result is related to lower gas production in 24 and 48 h at T-160. We concluded that the use of a blend of exogenous enzymes in the diet of lactating Jersey cows was able to increase milk production in these animals, resulting in greater feed efficiency and also an increase in milk protein content, positively modulating the fatty acid profile in the rumen and improving the apparent digestibility of nutrients. Full article

Defining appropriate dietary fiber levels is essential for enhancing the sustainability of feedlot lamb production. Optimal dietary fiber levels can enhance meat yield, improve nutrient retention and utilization, and reduce environmental impact. This study aimed to determine the optimal level of dietary fiber to enhance nutrient intake, digestibility, feeding behavior, and rumen fermentation in feedlot lambs. Five rumen-fistulated Santa Inês male lambs (40 kg, 7 months old) were used in a 5 × 5 Latin square design. Diets contained increasing levels of neutral detergent fiber (NDF): 200, 320, 440, 560, and 680 g/kg dry matter (DM), with each period lasting 21 days (total 105 days). Nutrient intake responded quadratically to NDF levels (p < 0.05). Apparent digestibility was significantly affected (p < 0.05), except for crude protein. Feeding (p = 0.001) and rumination times (p = 0.002) increased linearly, while idling time decreased (p < 0.001). Feeder visits declined (p = 0.002), and idling events followed a quadratic trend. Feeding and rumination efficiencies for DM decreased (p = 0.006 and p = 0.010), while NDF rumination efficiency increased (p = 0.014). The ruminal pH rose (p < 0.001), and propionate decreased (p = 0.019); acetate and butyrate showed quadratic responses. Based on intake, digestibility, and fermentation patterns, dietary NDF should be included at 400 g/kg DM to optimize nutrient utilization and rumen function in confined lambs. Full article

This study evaluated the effects of rumen-protected chromium-nicotinic acid (RPCNA) supplementation on lactation performance, nutrient digestion, ruminal fermentation, serum biochemical parameters and antioxidant in lactating water buffaloes under conditions of a critical thermal comfort index (TCI). Healthy lactating water buffaloes (milk yield = 5.96 ± 0.21 kg; parity = 2.96 ± 0.15, mean ± SD) were randomly assigned to four groups, which were the control (without RPCNA), and three treatment groups designated as RPCNA2, RPCNA4, and RPCNA6, with 0, 2, 4, and 6 mg/(d·head) of RPCNA, respectively. During a 56-day experimental period, the average temperature was 29 °C with an average TCI of 40.60, indicating a critical state of environmental stress. The results showed that the addition of RPCNA to the diet had no significant effect on the apparent digestibility of nutrients, lactation performance, and dry matter intake in lactating water buffaloes. However, RPCNA supplementation positively influenced rumen fermentation, reducing ammonia nitrogen (NH₃-N) concentrations and promoting microbial protein synthesis. Supplementation with RPCNA4 and RPCNA6 decreased rumen NH₃-N levels by 48.3% and 36.4%, respectively, while RPCNA4 increased isobutyrate concentrations. This demonstrates that the coating technology does not provide 100% rumen protection. Serum analysis revealed that RPCNA significantly increases total antioxidant capacity (T-AOC). Additionally, the supplementation of 4 mg/(d·head) of RPCNA, improved serum glutathione peroxidase (GSH-Px) activity and reduced malondialdehyde (MDA) levels. These findings suggest that moderate doses of RPCNA supplementation can improve antioxidant status and rumen nitrogen metabolism in lactating water buffaloes under critical state, without significantly altering milk production or composition. Full article

The effects of varying dietary metabolizable energy (ME) and crude protein (CP) levels, along with their interactive effects, on the apparent nutrient metabolism, development of digestive organs, intestinal morphology, and microbiota composition in Jingyuan chickens during the growing phase were evaluated. A total of 540 seven-week-old male Jingyuan chickens were randomly assigned to nine groups, with six replicates per group and 10 chickens per replicate. The trial lasted for 11 weeks. A 3 × 3 factorial design was adopted, comprising three levels of ME, namely, low (11.28 MJ/kg, LE group), medium (11.70 MJ/kg, ME group), and high (12.12 MJ/kg, HE group) and three levels of CP, namely, low (14.00%, LP group), medium (15.50%, MP group), and high (17.00%, HP group). The levels of ME and CP, along with their interactions, had significant effects on the average daily gain (ADG), average daily feed intake, feed conversion ratio (FCR), apparent metabolizable rate of CP, gizzard weight, duodenal lengths, jejunal villus height (VH), crypt depth (CD), and muscle layer thickness (MLT) (p < 0.05). The combination of medium level ME (11.70 MJ/kg) and medium level CP (15.50%) (MEMP group) exhibited the best performance, with the highest ADG and the lowest FCR (p < 0.05). Moreover, this group showed significantly higher duodenal length, jejunal CD, VH/CD and MLT compared with the other groups (p < 0.05). Dietary ME and CP levels greatly influenced cecal microbiota composition. Chickens in the MEMP group exhibited an increased abundance of Erysipelotrichaceae, Syntrophomonadaceae, Akkermansia, and Clostridia_vadinBB60_group, and there was an obvious decrease in the relative abundance of Desulfobacterota (p < 0.05). This study demonstrated that dietary ME and CP levels, along with their interactions, could significantly influence the growth performance, apparent nutrient metabolism, and intestinal development of Jingyuan chickens during the growing phase. Dietary ME and CP levels modulated the cecal microbiota composition, potentially inhibiting the abundance of harmful bacteria Desulfobacterota, while enriching the abundance of beneficial bacteria, thereby enhancing gut development and nutrient absorption. The combination of medium-level ME and CP (11.70 MJ/kg ME, 15.50% CP) demonstrated the most favorable outcomes in our study. Full article

The objective of this study was to investigate alterations in nutrient utilization, standardized ileal digestibility of amino acids (SIDAA), and intestinal health in response to heat stress (HS) in Pekin ducks. A total of 240 healthy 28-day-old male Pekin ducks were randomly allocated to three groups: a normal control (NC) group, an HS group, and a pair-fed (PF; provided an amount of feed equal to that consumed by the HS group to eliminate the effects of feed intake) group, each with eight replicate cages of ten birds. The results showed that HS significantly reduced the apparent utilization of dietary energy, ether extract, and crude protein compared to both the NC and PF groups (p < 0.05), but yielded comparable SIDAA to the PF group. The HS group exhibited reduced mRNA levels of EAAT3 and PepT1, along with elevated mRNA levels of CAT1, GLUT5, and FATP6 in the jejunum compared to the NC or PF groups, respectively (p < 0.05). Furthermore, HS resulted in a significant deterioration of jejunal morphology and goblet cell count compared to the NC and PF groups (p < 0.05). Serum fluorescein isothiocyanate-dextran levels were significantly higher in HS ducks than in NC ducks (p < 0.05), but did not differ from PF ducks. At order-level classification of ileal mucosal microbiota, HS markedly increased the relative abundance of Bacillales, Deferribacterales, and Actinomycetales versus NC (p < 0.05), while significantly decreasing Bifidobacteriales abundance relative to PF (p < 0.05). Collectively, HS induces a leaky gut and microbiota dysbiosis that compromises gut health, thereby reducing dietary nutrient utilization in Pekin ducks. The observed reduction in feed intake constitutes a primary driver of intestinal health deterioration in heat-stressed Pekin ducks. Full article

Our previous study has shown that supplementation of 0.50% benzoic acid (BA) increased growth performance, promoted rumen fermentation, and improved the composition and function of rumen microbiota. This research was designed to conduct a deeper exploration of the impacts of dietary supplementation with BA on the apparent digestibility of nutrients and the composition of rectal microbiota in weaned Holstein dairy calves. Sixteen Holstein heifer calves with similar body weights (91.2 ± 0.7 kg) were selected and randomly allocated into two groups, each comprising eight calves. Calves in the control group (CON group) were fed with a basal diet, while those in the benzoic acid group (BA group) were fed with the basal diet supplemented with 0.50% benzoic acid (on a dry matter basis). The experimental period started at 60 days of age and ended at 102 days of age, lasting for a total of 42 days. The calves were weaned at 60 days of age, with a transition period of 7 days. Feed samples were collected every two weeks, fecal samples were collected from 99 to 101 days of age, and blood samples were collected at 102 days of age. The results showed that supplementation with BA did not influence the digestibility of dry matter, crude protein, ether extract, neutral detergent fiber, acid detergent fiber, calcium, and phosphorus between the two groups. Compared with the CON group, BA supplementation tended to decrease the total cholesterol (TC) in the serum of the calves (p = 0.067). Supplementation with BA increased the relative abundances of the two beneficial bacteria, Bifidobacterium and Bifidobacterium pseudolongum (p < 0.05, LDA > 2), but decreased that of the harmful bacterium, Clostridium sensu stricto 1, in the rectum of dairy calves. The microbial functional prediction revealed that the fecal microbial metabolism involved in primary bile acid biosynthesis was higher in the calves from the BA group. In conclusion, the present study demonstrated that adding 0.50% BA to the diet did not influence the apparent nutrient digestibility, but improved rectal microbiota health, which finally promoted the growth performance in weaned Holstein dairy calves. Full article

Background: The current study aimed to evaluate the effects of the supplementation of protease sources on growth and carcass response, gut health, nutrient digestibility, and cecal microbiota profiles in broilers fed poultry-by-product-meal (PBM)-containing diets. Methods: In total, 800 one-day-old mixed-sex broilers (Arbor Acres) were weighed and allocated to one of the four dietary treatments in a completely randomized design, with eight replicates and 25 birds each per replicate. The treatments were as follows: (1) T₀, control diet (without protease supplementation and 3% PBM); (2) T₁, control diet supplemented with acidic protease at 100 g/ton (50,000 U/g); (3) T₂, control diet supplemented with alkaline protease at 200 g/ton (25,000 U/g); (4) T₃, control diet supplemented with neutral protease at 200 g/ton (25,000 U/g). Results: Protease supplementation enhanced (p < 0.05) body weight gain and the feed conversion ratio, predominantly in broilers fed PBM-based diets containing alkaline protease. Alkaline protease supplementation increased (p < 0.05) the apparent ileal digestibility of proteins (AIDP) by 4.3% and the apparent ileal digestibility of amino acids (AIDAA) by up to 5.8%, except for ornithine. Increments (p < 0.05) in carcass, breast, and leg quarter yields due to protease supplementation were evident, particularly in broilers fed diets containing alkaline protease. Alkaline protease improved (p < 0.05) the duodenal villus height (VH), reduced the crypt depth (CD), and increased the villus height to crypt depth ratio (VCR). Alkaline protease supplementation reduced (p < 0.05) cecal counts of Salmonella, Escherichia coli, and Clostridium in the broilers, whereas it increased (p < 0.05) the Lactobacillus counts. Conclusions: the supplemented alkaline protease resulted in improved growth performance and carcass traits, better gut health, as well as improved ileal digestibility of nutrients, including crude protein (CP) and acid insoluble ash (AIA), with a more balanced cecal microbial composition in broilers. Full article

This study used integrated in vitro and in vivo approaches to investigate how the starch source (glutinous rice, indica rice, maize, or high-amylose rice) influences starch digestion kinetics and, consequently, the apparent nutrient utilization and amino acid metabolism in goslings. Four diets were formulated using glutinous rice, indica rice, maize, and high-amylose rice, and in vitro digestion and animal experiments were carried out. The data showed the particle sizes of the four starches: glutinous rice ≈ indica rice < corn < amylose. The glutinous rice starch grain is a porous polyhedron with an angular surface, the corn starch grain is an ellipsoid with a smooth surface, the indica rice starch grain is a polyhedron with a smooth and compact surface, and the high-amylose starch grain is an irregular polyhedron with a smooth surface. Starch digestibility was relatively stable for the indica and corn-based diets, and starch digestibility was higher for the indica rice diet compared to the corn- and high-amylose starch-based diets. The utilization of Asp, Ser, Glu, Gly, and Phe was higher for the glutinous rice diet compared to the maize and high-amylose diets. Furthermore, with this diet, the availability of Thr and Ala was observed to be higher than with the indica rice and high-amylose diets. In conclusion, the particle size and structure of starch from different sources (glutinous rice, indica rice, corn, and high-amylose rice) were different, significantly affecting the starch digestion rate. The glutinous rice diet enables a fast digestion rate for starch, which is rapidly digested in the proximal intestine. The inadequate supply of glucose in the distal intestine enhances the oxidative energy supply from dietary amino acids in that region, thereby improving the apparent digestibility of both starch and crude protein. Full article

The apparent ileal digestibility (AID) and standardized ileal digestibility (SID) of amino acids, digestible energy (DE), metabolizable energy (ME), and the apparent total tract digestibility (ATTD) of nutrients in 10 fishmeal (FM) samples were evaluated in weaned barrows (Duroc × Landrace × Yorkshire) using two experiments. In Experiment 1, 11 piglets (18.87 ± 0.10 kg) fitted with T-cannulas were randomly allocated to an 11 × 6 Latin-square design with 11 diets (1 nitrogen-free diet and 10 assay diets) and six periods. The AID and SID of all amino acids (AAs) except proline showed significant differences among all FM (p < 0.05). Importantly, the SID of amino acids was positively correlated with key antioxidant markers and immune parameters, and it was negatively correlated with oxidative stress markers (MDA) and pro-inflammatory cytokines (IL-2 and IL-6). In Experiment 2, 11 piglets (18.05 ± 1.15 kg) were assigned to an 11 × 5 Latin-square design with 11 diets (a 96.35% corn diet and 10 assay diets) and five consecutive periods. Significant variations were observed in the DE, ME, and ATTD of dry matter among different FM samples (p < 0.05). Moreover, predictive equations for estimating the SID of lysine, methionine, threonine, and tryptophan, as well as DE and ME, were established using stepwise regression analysis based on the chemical composition of the FM. These findings demonstrate that the nutritional value of FM in nursery pig diets has been underestimated, and this study provides precise data and predictive methods for evaluating the nutritional quality of FM in precision nutrition. Full article

This study aimed to evaluate the effects of dietary nano-copper supplementation on growth performance, nutrient digestibility, antioxidant status, inflammatory response, and intestinal barrier function in weanling pigs under heat stress conditions. Forty 20-day-old weaned weanling pigs (Yorkshire × Landrace × Duroc) weighing 6.49 ± 0.08 kg were randomly divided into five treatments with eight replicates each. The pre-feeding period was 2 days, followed by a 22-day experimental period. All groups were exposed to high heat conditions at 35 ± 1 °C. The control group received a basal diet, while the low copper sulfate (LC) group received a diet with 50 mg/kg of copper sulfate, the high copper sulfate (HC) group received a diet with 150 mg/kg of copper sulfate, the low nano-copper (LNC) group received a diet with 50 mg/kg of nano-copper oxide, and the high nano-copper (HNC) group received a diet with 150 mg/kg of nano-copper oxide. Compared to the basal group, pigs supplemented with copper (either CuSO₄ or nano-CuO) exhibited significantly higher average daily gain (ADG, p < 0.048) and feed intake (ADFI, p = 0.005), with the 50 mg/kg nano-copper group showing improved nutrient digestibility (p < 0.05) and intestinal morphology. Nano-copper supplementation significantly enhanced mucosal SOD activity (p < 0.05), reduced MDA levels (p < 0.05), and downregulated pro-inflammatory cytokines such as IL-1β and IL-6 (p < 0.05). Notably, 50 mg/kg of nano-copper increased the apparent total tract digestibility (ATTD) of copper to 30.29%, significantly higher than the 16.55% observed in the 150 mg/kg CuSO₄ group (p < 0.05). Furthermore, fecal copper concentration was significantly reduced by 20.7% in the 50 mg/kg nano-copper group compared to copper sulfate (p < 0.001). In conclusion, nano-copper appears to be a promising alternative to copper sulfate for improving growth performance and reducing fecal copper concentrations in weanling pigs under heat stress conditions. Full article

Dairy cows are suffering from heat stress (HS) worldwide, and this has become a continual challenge in dairy production systems. The objective of this study was to investigate the effects of the shade of poplar trees on alleviating HS via measuring milk production, nutrient digestibility, immunity, and antioxidant capability in the serum of dairy cows in open cowsheds. A total of 540 lactating Holstein cows were assigned to 2 groups (3 cowsheds per group, 90 cows per cowshed), including the treatment group with poplar trees on the west side of sheds and the control group without trees. This study was carried out in the early-hot season, hot season, and late-hot season. The results showed the following: (1) During the hot season, shaded cows exhibited lower respiration rate (p < 0.05) and higher dry matter intake (p < 0.05) and daily milk yield (p < 0.05), compared with the control. Also, apparent digestibility of crude protein, acid detergent fiber, and neutral detergent fiber demonstrated an increase (p < 0.05) in shaded cows during the hot season. (2) Total antioxidant capacity and superoxide dismutase activity in the serum of shaded cows increased (p < 0.05) during the hot season and late-hot season, compared with the control. (3) Serum interleukin-4, immunoglobulin G, and immunoglobulin M concentrations of shaded cows were greater (p < 0.05) than the control during the hot season. Additionally, serum concentrations of HSP60, HSP70, and HSP90 in shaded cows showed a decrease (p < 0.05) during the hot season. In conclusion, shade from poplar trees can mitigate the adverse impact of high-temperature environments on cows in open cowsheds. Full article

This study aimed to evaluate the apparent digestibility coefficients (ADCs) for nutrients and energy of seven conventional and alternative feed ingredients (poultry feather meal, fermented feather meal, mealworm meal, defatted black soldier fly, Chlorella, poultry by-product meal, and corn meal) when fed to Sobaity seabream (Sparidentex hasta), with the goal of identifying sustainable, digestible, and nutritionally viable ingredients for aquaculture feed formulations. A reference diet (RF) was formulated to meet the nutrient requirements of Sobaity seabream while test diets were prepared to contain 70% RF and 30% of the test ingredients. Sobaity seabream (200 ± 8.0 g) were fed the diets for seven days before fecal matter was collected by stripping. The whole length of the digestibility trial was 21 days. The ingredient apparent digestibility of dry matter (34.8–70.4%), crude protein (52.8–107.8%), crude lipid (67.7–112.9%), and energy (52.2–86.1%) were affected by test ingredients (p < 0.01). The dry matter digestibility of mealworm meal was the highest (70.4%) compared to other ingredients. Feather meal, Chlorella, and black soldier fly meal had significantly lower values of dry matter digestibility. Dry matter and crude protein were significantly more digestible in fermented feather meal than the feather meal without fermentation. The crude protein digestibility was significantly higher (107.8%) for mealworm meal. However, feather meal has shown a significantly lower value (52.8%) for crude protein digestibility compared to other ingredients. Energy digestibility showed a significant positive correlation with dry matter digestibility (r = 0.870). The energy digestibility of mealworm meal was significantly higher (86.1%, p < 0.05) than other ingredients. Feather meal had the lowest energy digestibility (52.2%) with no statistically significant difference from Chlorella, corn meal, and black soldier fly meal. This study indicates that mealworm meal is the most easily digestible protein source for Sobaity seabream and should be prioritized in their diets. Fermentation enhances the digestibility of feather meal and is recommended when using it. Ingredients with a lower digestibility, like feather meal, chlorella, and black soldier fly meal, should be used in moderation or undergo further processing to improve nutrient availability. Full article

The purpose of this research was to investigate the impact of chestnut tannins (CHTs) on nutrient digestibility, nitrogen balance, in situ crude protein (CP) digestibility, protease enzymes, and microbial community composition in sheep. Eighteen 1.5-year-old sheep (43.0 ± 2.0 kg initial BW) fitted with permanent ruminal cannula were selected and randomly divided into three groups, which were fed with CHTs added at 0, 2, and 6%/kg DM. The pre-feeding period lasted for 12 days, and the actual trial period was 18 days. Rumen fluid was collected to assess in situ crude protein (CP) degradability, while rumen digesta was analyzed for total and ruminal proteolytic bacterial populations. Using one-way ANOVA in SAS to analyze data, the results indicated that 2% CHT reduced in situ degradability by 26.23%, while 6% reduced it by 58.01% in the rumen of the sheep. The CP apparent digestibility, nitrogen metabolism, and population of proteolytic bacteria of sheep were decreased in the 6% CHT group (p < 0.05), while the above indices of the 2% CHT group were not affected. Furthermore, CHT supplementation significantly altered the ruminal microbial community structure. Particularly in the 2% CHT group, the relative abundances of Bacteroidota and Prevotella increased. LEfSe analysis revealed that Bacteroidale replaced U29-B03 as the dominant microbiota at 2% CHT. Doses of 2% CHT can be incorporated into sheep diets without impairing digestion. These findings support the inclusion of CHT doses of less than 2% for enhancing protein digestion and increasing the types of beneficial bacteria in the rumen, while doses above 6% should be avoided. Full article