The inclusion of fibrolytic enzymes in the diet is believed to have positive effects on animal production. Hence, the objective of this study was to investigate the impact of supplementing diets with a commercial fibrolytic enzyme preparation (Vistamax; mixture of xylanase and cellulase) derived from Trichoderma reesei on lactational performance, digestibility, and plasma nutrient levels in high-producing dairy cows. Two dietary energy levels were considered: a normal energy diet (metabolizable energy = 2.68 Mcal/kg) and a slightly lower energy diet (metabolizable energy = 2.55 Mcal/kg). A total of 120 lactating Holstein cows (parity = 2; Days in Milk = 113 ± 23) were randomly assigned to four treatment groups using a 2 * 2 factorial arrangement. The dietary treatments consisted of: (1) normal energy diet without enzyme supplementation (NL); (2) normal energy diet with enzyme supplementation (NLE); (3) slightly lower energy diet without enzyme supplementation (SL); and (4) slightly lower energy diet with enzyme supplementation (SLE). The amount of enzyme added to the diets was determined based on previous in vitro studies and supplier recommendations. The enzyme and premix were mixed prior to the preparation of the total mixed ration, and the trial lasted for a duration of 42 days. The results indicated that the application of the fibrolytic enzyme did not have a significant effect on dry matter intake (DMI), but it did enhance the digestibility of dry matter (DM), neutral detergent fiber (NDF), potentially digestible NDF (pdNDF), organic matter (OM), milk production, milk urea nitrogen (MUN), and blood urea nitrogen (BUN). On the other hand, the slightly lower energy diet resulted in a decrease in DMI, milk production, milk protein yield, plasma free amino acids (FAA), and an increase in plasma B-hydroxybutyrate (BHBA). In conclusion, the inclusion of the fibrolytic enzyme in the diets of dairy cows led to improvements in the digestibility of DM, NDF, pdNDF, OM, milk production, and feed efficiency. Furthermore, the application of the enzyme to the slightly lower energy diet resulted in milk production levels comparable to those observed in cows fed the untreated normal energy diet.

This study was to investigate the effects of cellulase and xylanase on fermentation characteristics, nutrient composition and the bacterial community of the mixed silage of king grass and rice straw. Lab-scale bag silage was produced and seven groups were studied: blank control (CK); added 1%, 2% cellulase (CE1, CE2); added 1%, 2% xylanase (XY1, XY2); and added 0.5% cellulase +0.5% xylanase, 1% cellulase +1% xylanase (CX1, CX2). The results showed that the application of additives in six treated groups exerted a positive effect on lactic acid (LA) content and their pH values decreased significantly (p < 0.05). The addition of cellulase and xylanase decreased (p < 0.05) the content of neutral detergent fiber (NDF) and acid detergent fiber (ADF) significantly and increased (p < 0.01) the crude protein (CP) and water-soluble carbohydrate (WSC) content. Filter paper enzyme activity (FPA) declined and xylanase activity (XA) intensified (p < 0.05) as ensiling was prolonged, where most of the enzymatic treatments (especially XY2, CX2) resulted in increased enzyme activities. Moreover, the addition of cellulase and xylanase reduced the abundance of harmful bacteria such as Acinetobacter and Klebsiella and increased the abundance of lactic acid bacteria such as Lacticaseseibacillus, Lactiplantibacillus. In conclusion, the addition of cellulase and xylanase would improve fermentation quality and nutrient preservation via altering the bacterial community, with 1% cellulase or complex enzyme best.

The present experiment evaluated the feeding of date palm leaves (DPL) ensiled with fibrolytic enzymes (ENZ) or multi-species probiotics (MSP) on nutrient utilization and lactational performance of ewes. Fifty multiparous lactating Farafra ewes were used in a completely randomized design for 90 d. The treatments consisted of the control diet with a concentrate feed mixture and date palm leaves (at 60:40, DM basis) ensiled without additive (control) or DPL ensiled with ENZ or MSP replacing control DPL at 50 or 100%. Both ENZ and MSP increased (p < 0.01) DPL and total intakes, digestibility of all nutrients, concentrations of ammonia, total volatile fatty acids, acetate and propionate in the rumen. Increased milk production, concentrations of fat, lactose and energy in milk, and feed efficiency were observed with MSP and ENZ compared to the control treatment. Moreover, ENZ and MSP increased (p < 0.05) the concentrations of total n3, n6 fatty acids, polyunsaturated fatty acids and conjugated linoleic acids and decreased (p < 0.001) the atherogenicity. The differences between ENZ and MSP and between the low and high replacement levels were minor for all measured parameters. Ensiling of DPL with MSP or fibrolytic enzymes is recommended to improve feed efficiency and improve lactational performance of ewes.

This study was conducted to evaluate the effects of adding different levels of the combination of fibrolytic enzymes and probiotics (a mixture of bacteria and yeast) on the performance of fattening lambs. Thirty-two male Ossimi lambs (weighing 39 ± 0.24 kg) were divided into four groups randomly (eight animals each). The first group (control ration, G1) was fed on a ration of 60% concentrate feed mixture (CFM), 20% Egyptian clover (EC), and 20% wheat straw (WS). The second (G2), third (G3), and fourth (G4) groups were fed a control ration supplemented with Calfo Care® at concentrations of 0.5, 1, and 2 kg/ton diet of dry matter (DM). Results showed that the G2 and G3 rations significantly (p ≤ 0.05) increased the DM, organic matter, crude protein, crude fiber, and ether extract digestibility compared with the G1 and G4 rations. Moreover, the G2 and G3 rations increased (p ≤ 0.05) the percentages of total digestible nutrients (TDN), starch values (SV), and digestible crude protein (DCP) compared with the G1 and G4 rations. Both the G2 and G3 rations significantly (p ≤ 0.05) increased the TDN, SV, and DCP as kg/day or g/kg w0.75 and kg or g/100 kg body weight compared with the G1 and G4 rations. Conversely, the G1 ration significantly decreased the feed conversion of DM, TDN, SV, and DCP compared with the experimental groups. Furthermore, the G2, G3, and G4 rations significantly (p ≤ 0.05) increased the total weight gain by 25.34%, 52.20%, and 3.79%, respectively, compared with the G1 ration. The G2, G3, and G4 rations also (p ≤ 0.05) increased the concentrations of most hematological parameters, including triiodothyronine, total protein, albumin, and glucose, compared with the G1 ration. Finally, the best net profit was recorded with the G3 ration, followed by the G2, G4, and G1 rations.

The fibrolytic enzymes and the hindgut fungi in donkey cecum-colon ecosystem play an important role in dietary fiber digestion. A better understanding of the fibrolytic enzyme profiles and the fungal community along donkey caecum and colon is key for optimizing hindgut function. In the present study, the fibrolytic enzyme activities within donkey caecum and colon were firstly measured by spectrophotometry. Activities of carboxymethyl cellulase, avicelase, xylanase, and acetyl esterase were greater in donkey dorsal colon than in caecum, indicating that the colon microorganisms may be more efficient in producing fibrolytic enzymes compared to caecum microbes. The fungal community composition along donkey hindgut was determined by sequencing ITS region using Illumina MiSeq. Three fungal phyla were identified by sequence comparison: Ascomycota (66.8%-74.4%), Basidiomycota (21.6%-30.9%), and Neocallimastigomycota (0.9%-3.3%). The Aspergillus, Wallemia, Phanerochaete, Fusarium, and Penicillium were detected as the dominant genera, but their metabolic and functional significance in donkey cecum-colon ecosystem need further investigation. In terms of the anaerobic fungi Neocallimastigomycota, its abundance was greater in donkey colon than in caecum (p < 0.05), indicating that the donkey hindgut region was associated with differences in fungal community composition. Moreover, the relative abundance of enzymes related to plant cell wall degradation were predicted by PICRUSt, and they were also lower in caecum than in colon. The present study provided new information about fibrolytic enzyme profiles and fungal composition in donkey hindgut ecosystem.

The objective of this study was to examine the enzyme activities of an enzymatic complex produced by Pleurotus ostreatus in different pH and the effects of adding increased application rates of this enzymatic complex on the fermentation profile, chemical composition, and in situ ruminal disappearance of whole-plant corn silage (WPCS) at the onset of fermentation and 30 d after ensiling. The lignocellulolytic enzymatic complex was obtained through in vitro cultivation of P. ostreatus. In the first experiment, the activities of laccase, lignin peroxidase (LiP), manganese peroxidase, endo- and exo-glucanase, xylanase, and mannanase were determined at pH 3, 4, 5, and 6. In the second experiment, five application rates of enzymatic complex were tested in a randomized complete block design (0, 9, 18, 27, and 36 mg of lignocellulosic enzymes/kg of fresh whole-plant corn [WPC], corresponding to 0, 0.587, 1.156, 1.734, and 2.312 g of enzymatic complex/kg of fresh WPC, respectively). There were four replicates per treatment (vacuum-sealed bags) per opening time. Bags were opened 1, 2, 3, and 7 d after ensiling (onset of fermentation period) and 30 d after ensiling to evaluate the fermentation profile, chemical composition, and in situ dry matter and neutral fiber detergent disappearance of WPCS. Laccase had the greatest activity at pH 5 (P < 0.01), whereas manganese peroxidase and LiP had the greatest activity at pH 4 (P < 0.01; P < 0.01). There was no effect of the rate of application of enzymatic complex, at the onset of fermentation, on the fermentation profile (P > 0.21), and chemical composition (P > 0.36). The concentration of water-soluble carbohydrate quadratically decreased (P < 0.01) over the ensiling time at the onset of fermentation, leading to a quadratic increase of lactic acid (P = 0.02) and a linear increase of acetic acid (P = 0.02) throughout fermentation. Consequently, pH quadratically decreased (P < 0.01). Lignin concentration linearly decreased (P = 0.04) with the enzymatic complex application rates at 30 d of storage; however, other nutrients and fermentation profiles did not change (P > 0.11) with the enzymatic complex application rates. Addition of lignocellulolytic enzymatic complex from P. ostreatus cultivation to WPC at ensiling decreased WPCS lignin concentration 30 d after ensiling; however, it was not sufficient to improve in situ disappearance of fiber and dry matter.

The objective of this study was to evaluate the effects of exogenous enzymes on nutrient intake and digestibility, rumen fermentation, and productivity of mid-lactating cows. Experiment 1 was designed to test increasing doses [0, 0.5, 1.0, or 1.5 g/kg of dry matter (DM)] of a combination of 2 enzyme products with xylanase and β-glucanase activities (Ronozyme Wx and Ronozyme VP, respectively; DSM Nutritional Products) on rumen fermentation and total apparent digestibility. Enzyme combinations had a ratio of endo-1,3(4)-β-glucanase to endo-1,4-β-xylanase of 8:2 (wt/wt). For experiment 1, 8 rumen cannulated lactating cows were used into a double 4 × 4 Latin square design experiment with 14 d of diet adaptation and 7 d of sampling. Despite no differences in feed intake, carbohydrases linearly increased neutral detergent fiber digestibility. Treatments marginally affected rumen fermentation, where a linear trend for lower rumen pH and a linear trend for greater isobutyrate concentration were observed with increasing enzyme dose. A trend for lower rumen NH3-N concentration was observed for cows receiving carbohydrases in comparison with control group. When comparing all enzyme treatments against control group, cows fed enzymes tended to produce more 3.5% fat-corrected milk (FCM), produced more milk fat, and had greater blood glucose concentration. Experiment 2 evaluated 3 doses (0, 0.5, or 0.75 g/kg of DM) of the same combination of enzyme products on performance of cows (n = 36) in a complete randomized block (n = 12) design. Cows received treatments for 9 wk. No interaction effects between treatments and time were observed for all variables assessed in this study. In agreement with experiment 1, no differences were detected for feed intake, but cows fed the enzyme products tended to produce more 3.5% FCM and milk fat compared with control. In addition, cows fed enzymes exhibited greater efficiency of FCM production (FCM ÷ DM intake) compared with control. No differences were detected for intake and productivity when comparing the 2 doses of carbohydrases. In summary, the enzyme products tested in this study may improve feed efficiency due to greater milk fat concentration.

Pleurotus ostreatus (oyster mushroom) synthesizes enzymes that degrade lignin, cellulose, and hemicellulose. The objectives of this study were to evaluate the effect of Pleurotus ostreatus spent substrate (POSS) on whole-plant corn silage (WPCS) chemical composition, antioxidant capacity, lignin monomers, and in vitro digestibility, as well as the performance of lactating goats fed corn silage treated with different levels of POSS. In experiment 1, 4 levels of lignocellulolytic enzymes were tested in a complete randomized design: 0, 10, 20, and 30 mg of lignocellulosic enzymes per kilogram of fresh matter, 4 replicates per treatment (vacuum-sealed bags). The bags were opened 60 d after ensiling. In experiment 2, corn silage treated with 3 enzyme levels (0, 10, or 30 mg/kg of fresh matter) was fed to lactating goats as part of the total mixed ration. Nine lactating Saanen goats (62.68 ± 7.62 kg BW; 44 ± 8 d in milk; 2.91 ± 0.81 kg of milk/day, mean ± SD) were assigned to three 3 × 3 Latin squares. Data were analyzed using the GLIMMIX procedure of SAS (version 9.4, SAS Institute Inc.), and means were compared by linear and quadratic orthogonal contrast. In experiment 1, neutral detergent fiber (NDF), acid detergent fiber (ADF), lignin, and cellulose quadratically decreased in the WPCS treated with POSS. At the nadir point, POSS decreased NDF by 14.1%, ADF by 19.5%, lignin by 9.07%, and cellulose by 22.1% compared with the untreated silage. Therefore, POSS led to a quadratic increase in in vitro dry matter digestibility of WPCS (+8.88% at the vertex) compared with the untreated silage. In experiment 2, POSS quadratically increased the in vivo total-tract ADF digestibility. Also, the concentration of polyphenols in the milk of goats linearly increased with the addition of POSS, and no differences were observed among treatments for milk yield and composition. In summary, adding 10 mg of lignocellulolytic enzymes from POSS per kilogram of fresh matter of whole-plant corn at ensiling had a more evident reduction in lignin and cellulose concentration, leading to greater in vitro digestibility, as well as greater in vivo ADF digestibility; however, milk yield was not different among treatments.

This study\'s objectives were to (a) consider the combined effects of Lactobacillus casei TH14 and additives on fermentation quality, nutritive value and microbial composition when ensiling rice straw and (b) evaluate the ensiled rice straw using the in vitro gas production technique. A 2 × 2 × 2 factorial arrangement in a completely randomized design (CRD) was used in a randomized complete design with three replications. The rice straw was treated with L. casei TH14 (T) at 0 and 105 colony-forming unit (cfu)/kg fresh matter (FM), cellulase (C) at 0 and 10,000 unit/kg FM and molasses (M) at 0 and 50 g/kg FM. After 30 days, the dry matter content of the ensiled rice straw was a range between 261.77 and 287.01 g/kg while untreated rice straw was 934.00 g/kg. Neutral detergent fibre (NDF) and acid detergent fibre (ADF) contents were decreased when treated with C (p < .01). Th e pH, ammonia-nitrogen, lactic acid, acetic acid and butyric acid values were affected by the interaction among the additives. In particular, pH was significantly (p < .01) lower in rice straw treated with T × C × M. Lactic acid bacterium (LAB) counts were highest when ensilage rice straw with T × C × M (p < .05), while untreated rice straw indicated highest amount for aerobic bacteria (p < .01) and yeast populations (p < .01). Interaction among additives had no effect (p > .05) on gas production rate constant for the insoluble fraction (c value), but combination between C × M with ensiled rice straw reduced the c value (p < .05). The in vitro dry matter digestibility at 48 hr after incubation increased when rice straw was treated with T × C (p < .01), T × M (p < .05) and C × M (p < .05). Total volatile fatty acids at 4 hr and the mean value were increased (p < .05) when ensiled rice straw with T × C or T × M. Propionic acid was improved by adding C or M (p < .05). T × M did not influence (p > .05) any ruminal microorganism counts except for bacterial counts at 4 hr (p < .01). Thus, ensiled rice straw with T × C × M could promote the quality of ensiled rice straw and improve gas kinetics, fibre digestion and ruminal characteristics.

Four experiments were conducted to examine the effects of a recombinant bacterial expansin-like protein (BsEXLX1) from Bacillus subtilis and a commercial exogenous fibrolytic enzyme (EFE) preparation for ruminants on hydrolysis of pure substrates (cellulose and xylan) and in vitro digestibility of bermudagrass haylage (BMH). Recombinant Escherichia coli BL21 strain was used to express BsEXLX1; the protein was purified using an affinity column. In experiment 1, carboxymethylcellulose, Whatman #1 filter paper (General Electric, Boston, MA) and oat-spelt xylan substrates were subjected to 4 treatments (1) sodium citrate buffer (control), (2) BsEXLX1 (162 µg/g of substrate), (3) EFE (2.3 mg/g of substrate), and (4) EFE + BsELX1 in 3 independent runs. Samples were incubated at optimal conditions for both additives (pH 5 and 50°C) or at ruminal (pH 6 and 39°C) or ambient (pH 6 and 25°C) conditions for 24 h and sugar release was measured. In experiment 2, digestibility in vitro of BMH was examined after treatment with the following: (1) control (buffer only), (2) BsEXLX1 (162 µg/g of dry matter), (3) EFE (2.2 mg/g of dry matter), and (4) EFE + BsEXLX1 in 3 independent runs at 39°C for 24 h. Experiment 3 examined effects of EFE and BsEXLX1 on simulated preingestive hydrolysis and profile of released sugars from BMH after samples were suspended in deionized water with sodium azide at 25°C for 24 h in 2 independent runs. In experiment 4, the sequence of the BsEXLX1 purified protein was compared with 447 ruminal bacterial genomes to identify similar proteins from the rumen. In experiment 1, compared with EFE alone, EFE and BsEXLX1 synergistically increased sugar release from carboxymethylcellulose and Whatman #1 filter paper under all simulated conditions; however, hydrolysis of xylan was not improved. In experiment 2, compared with EFE alone, treatment with EFE and BsEXLX1 increased neutral detergent fiber and acid detergent fiber digestibility of bermudagrass haylage (by 5.5 and 15%, respectively) and total volatile fatty acid concentrations, and decreased acetate-propionate ratio. In experiment 3, compared with EFE alone. The EFE and BsEXLX1 synergistically reduced concentrations of neutral detergent fiber and acid detergent fiber and increased release of sugars by 9.3%, particularly cellobiose (72.5%). In experiment 4, a similar sequence to that of BsEXLX1 was identified in Bacillus licheniformis, and similar hypothetical protein sequences were identified in Ruminococcus flavefaciens strains along with different protein structures in E. xylanophilum and Lachnospiraceae. This study showed that an expansin-like protein synergistically increased the hydrolysis of pure cellulose substrates and the hydrolysis and digestibility in vitro of BMH.