Compositions and methods for increasing utilization of non-digestible fibers in animals

Abstract

The present disclosure provides compositions and methods for increasing the utilization of non-digestible fibers (NDFs) in animal feed and reducing the anti-nutritional effects of the NDFs. The method comprises orally administering to the animal a composition comprising lysozyme. More specifically, the method comprises orally administering to the animal a composition comprising a lysozyme, i.e., a C-type lysozyme or a GH-22 lysozyme. The methods and compositions can be used to increase NDF utilization, growth performance, survival rate, or any combination thereof in the non-human animal.

Description

[0001] Cross-referencing related applications

[0002] This application claims the benefit of U.S. Provisional Application Serial No. 63 / 477,548, filed December 28, 2022, pursuant to 35 USC §119(e), the entire disclosure of which is incorporated herein by reference.

[0003] Background Technology and Summary of the Invention

[0004] Insoluble fiber components in feed have a negative impact on animal gut health. Previous studies have shown that feeding insoluble fiber increases the incidence and severity of swine dysentery by (1) increasing the production of unfavorable types of mucin and (2) reducing the abundance of lactobacillus bacteria, which play a significantly beneficial role in the gut of piglets with intestinal infections (see Li et al., 2019; Wilberts et al., 2014). In addition, insoluble dietary fiber reduces digestive transit time and also negatively impacts the ileal digestibility of amino acids and other nutrients. Improving the digestibility and utilization of insoluble fiber in feed components may also benefit the digestibility of other nutrients in the diet.

[0005] The fiber components in maize and maize DDGS are predominantly insoluble and poorly fermentable, with arabinosylxylan and cellulose being the main non-starch polysaccharides (Ranathunga et al., 2018). Previous studies have shown that feeding insoluble fiber from DDGS exacerbates E. coli infection in piglets, as measured by increased fecal scores and E. coli shedding (Li et al., 2019). One nutritional intervention to improve the digestibility of fiber, particularly the insoluble fiber component, is the addition of carbohydrate enzymes (e.g., xylanase and β-glucanase) to the diets of non-ruminants. However, responses to survival and performance have been highly inconsistent. For example, Petry and Patience (2022) reviewed 19 publications from 2000 to 2020 and reported that xylanase improved ADG and G:F in 33% and 26% of studies, respectively, but did not report improvements in average daily feed intake. The responses of xylanase to apparent ileal digestibility (AID) and apparent total tract digestibility (ATTD) of fiber (NSP, TDF, or NDF) also vary considerably. Xylanase improved fiber AID and ATTD by 61% and 50% of the time, respectively. Therefore, a feed additive is needed to improve utilization and reduce the anti-nutritional effects of poorly digestible fiber in animal feed.

[0006] Accordingly, the present disclosure provides such compositions and methods. The methods of the present disclosure can comprise orally administering to an animal a composition comprising a lysozyme, such as a C-type lysozyme or a GH-22 lysozyme. As described herein, the methods and the compositions can be used to increase NDF utilization, growth performance, survival rate, or any combination thereof in the non-human animal.

[0007] Other objects, features and advantages of the present disclosure will become apparent from the following detailed description. It is to be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the application, are given by way of illustration only, since various changes and modifications within the spirit and scope of the application will become apparent to those skilled in the art from this detailed description. BRIEF DESCRIPTION OF DRAWINGS

[0008] The detailed description refers to the accompanying drawings that show, by way of example:

[0009] Figure 1 A graph showing the effect of dietary supplementation of lysozyme on apparent total gut digestibility (ATTD) of dry matter in nursery pigs.

[0010] Figure 2 A graph showing the effect of dietary supplementation of lysozyme on apparent total gut digestibility (ATTD) of gross energy (GE) in nursery pigs.

[0011] Figure 3 A graph showing the effect of dietary supplementation of lysozyme on apparent total gut digestibility (ATTD) of nitrogen in nursery pigs.

[0012] Figure 4 A graph showing the effect of dietary supplementation of lysozyme on ADG in nursery pigs.

[0013] Figure 5 A graph showing a meta-analysis for ADG of feeding lysozyme to nursery pigs during early nursery period.

[0014] Figure 6 A graph showing a meta-analysis for ADFI of feeding lysozyme to nursery pigs during early nursery period.

[0015] Figure 7 A graph showing a meta-analysis for F:G of feeding lysozyme to nursery pigs during early nursery period.

[0016] Figure 8 A graph showing a meta-analysis for BW of feeding lysozyme to nursery pigs during early nursery period.

[0017] Figure 9 A graph showing a meta-analysis for total removal rate of feeding lysozyme to nursery pigs during early nursery period.

[0018] Figure 10 A graph showing a meta-analysis of feeding lysozyme to nursery pigs during the early nursery period for mortality is shown.

[0019] Figure 11 A graph showing a meta-analysis of feeding lysozyme to nursery pigs throughout the nursery period (49-51 days post-weaning) for finish nursery performance is shown.

[0020] Figure 12 A graph showing a meta-analysis of feeding lysozyme to nursery pigs throughout the nursery period (49-51 days post-weaning) for finish nursery performance is shown. DETAILED DESCRIPTION

[0021] Feed composition

[0022] One aspect of the present disclosure encompasses a feed composition for a non-human animal. The composition comprises a basal animal diet supplemented with lysozyme. Lysozyme is a bacteriolytic enzyme that hydrolyzes the beta (1->4) bond between N-acetylglucosamine and N-acetylmuramic acid in the peptidoglycan of prokaryotic cell walls. It is also recruited for digestion in certain ruminants and colobine monkeys. There are at least five different classes of lysozymes: C (chick-type; C-type lysozyme), G (goose-type), phage-type (T4), fungal (Chalaropsis; CH-type lysozyme), and bacterial (Bacillus subtilis). The sequences of the different types of lysozymes have little in common.

[0023] In some aspects, the lysozyme of the present disclosure is a C-type lysozyme. C-type lysozymes belong to the glycoside hydrolase family 22 (GH-22 lysozyme). Thus, in some aspects, the lysozyme of the present disclosure is a GH-22 lysozyme. In some aspects, the lysozyme of the present disclosure is a C-type lysozyme. In some aspects, the lysozyme of the present disclosure is an egg white lysozyme. In some aspects, the concentration of egg white lysozyme in the feed composition ranges from about 0.1 lb / ton to about 0.5 lb / ton. In some aspects, the concentration of egg white lysozyme in the feed composition ranges from about 0.2 lb / ton to about 0.4 lb / ton. In some aspects, the concentration of egg white lysozyme ranges from about 0.001% w / w to about 0.5% w / w. In some aspects, the concentration of egg white lysozyme ranges from about 0.005% w / w to about 0.250% w / w.

[0024] In some aspects, the lysozyme of the present disclosure is not a CH-type lysozyme. CH-type lysozymes belong to the glycoside hydrolase family 25 (GH-25 lysozyme). Thus, the lysozyme of the present disclosure is not a GH-25 lysozyme.

[0025] Feed additive composition

[0026] One aspect of the present disclosure encompasses a feed additive composition for a non-human animal. The feed additive composition comprises a lysozyme. The lysozyme can be as described herein above in Section I. In addition to the lysozyme, the feed additive composition of the present disclosure can further comprise at least one additional ingredient such as a vitamin, a mineral, an amino acid, an antioxidant, a probiotic, an essential fatty acid, and a pharmaceutically acceptable excipient. The feed additive composition can be added to a basal animal diet for administration to a non-human animal. The feed additive composition can be formulated with a basal animal diet to make the feed composition previously described.

[0027] The amount of lysozyme in the feed additive composition can and will vary depending on the lysozyme, the type of non-human animal to which the feed additive composition comprising the lysozyme will be administered, the body weight, sex, and medical condition of the non-human animal to which the feed additive composition will be administered. Generally, the feed additive composition can comprise from about 20% to about 95% of the lysozyme.

[0028] In various aspects, the feed additive composition can be introduced into the basal animal diet by various methods depending on whether the feed additive composition is in liquid or solid form. Non-limiting examples of introducing the feed additive composition into the basal animal diet can be formulating the feed additive composition into the basal animal diet, topdressing the solid composition in the basal animal diet, spraying the liquid feed additive composition onto the basal animal diet, or combinations thereof. It should be recognized that when the feed additive is introduced into the basal animal diet, the amount of the feed additive introduced into the basal animal diet is sufficient to provide a therapeutically effective amount of the combination of the formulated yeast and pepper product in the diet of the animal.

[0029] Method of use

[0030] Another aspect of the present disclosure encompasses a method of nutritional intervention using dietary supplementation with a lysozyme. The method comprises orally administering an animal feed composition or a feed additive of the present disclosure to a non-human animal. The feed composition can be utilized as previously described, and the feed additive composition can be utilized as previously described.

[0031] In broad terms, a non-human animal can be defined as any animal that exhibits improved growth, improved health, improved gut health, and reduced microbial pathogen counts following administration of a feed additive composition. In various aspects, the non-human animal can be a domestic mammal of varying age and health status. Non-limiting examples of suitable domestic mammals can be beef cattle, horses, dairy cattle, calves, swine, goats, sheep, bison, llamas, or alpacas. In other aspects, the non-human animal can be a bird species of varying age and health status. Non-limiting examples of suitable bird species or poultry can be chickens (including broilers, layers, and breeders), ducks, pullets, geese, pheasants, guinea fowl / hens, quail, turkeys, and ratites such as emus and ostriches, and aquaculture. In another aspect, the non-human animal can be a companion animal of varying age and health status. Non-limiting examples of companion animals can be dogs, cats, birds, hamsters, or guinea pigs. In some aspects, the non-human animal is selected from the group comprising growing pigs, calves, foals, lambs (goats), kids, crias, chicks, young birds, ducklings, puppies, kittens, or combinations thereof.

[0032] In some aspects, the non-human animal is a non-ruminant animal. In some aspects, the non-human animal is a pig. In some aspects, the non-human animal is a nursery pig. In some aspects, the non-human animal is a healthy nursery pig.

[0033] The methods can be used to improve NDF utilization in animal feed. The methods of the present disclosure can also be used to reduce the anti-nutritional effects of indigestible fiber in animal feed. In some aspects, the methods of the present disclosure comprise reducing the incidence and severity of pig scour, reducing the production of undesirable types of mucin, increasing the abundance of lactic acid-producing bacteria (i.e., lactobacilli), increasing digesta transit time, positively affecting ileal digestibility of amino acids and other nutrients, or any combination thereof. The methods can improve nutrient digestibility, fiber digestibility, and growth performance of an animal without affecting E. coli proliferation.

[0034] In some aspects, the methods increase NDF digestibility by at least 36%. In some aspects, the methods reduce undigested NDF of DDGS by at least 13%. The methods can increase NDF utilization in the presence or absence of a pathogen.

[0035] In some aspects, the methods increase nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof. For example, the methods can increase ADG, ADFI, feed conversion, increase body weight, decrease mortality, decrease total removal, or any combination thereof. The methods can also increase apparent total digestive tract digestibility (ATTD) of dry matter, increase ATTD of total energy, increase ATTD of nitrogen. In some aspects, the methods reduce the indigestible carbohydrate fraction in the feed composition.

[0036] In some aspects, the methods of the present disclosure comprise improving growth performance, survivability, or growth performance and survivability of a non-human animal. The methods comprise orally administering to the animal a feed composition or a feed additive composition comprising lysozyme. The lysozyme, feed composition, and feed additive composition can be as previously described.

[0037] In some aspects, the methods improve growth performance, survivability, or growth performance and survivability of the animal regardless of the disease status of the animal. In other aspects, the methods improve nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof. In some aspects, the methods improve ADG, ADFI, feed conversion, increase body weight, decrease mortality, decrease total removal, or any combination thereof. In other aspects, the methods improve apparent total digestive tract digestibility (ATTD) of dry matter, improve ATTD of total energy, improve ATTD of nitrogen. In further aspects, the methods improve growth performance, survivability, or growth performance and survivability without affecting the proliferation of a pathogen in the animal. The methods can also reduce the indigestible carbohydrate fraction in the feed composition.

[0038] The time and duration of administration of the compositions of the present disclosure to an animal can and will vary. The feed or feed additive composition can be administered throughout the period that the animal is being fed. Alternatively, the feed or feed additive compositions of the present disclosure can be administered at specific periods during the growth and development of the animal. The feed or feed additive compositions of the present disclosure can also be administered at various intervals. For example, the compositions can be administered daily, weekly, monthly, or over a period of months. In some aspects, the compositions are administered weekly. In other aspects, the compositions are administered monthly. In some aspects, the compositions are administered daily. As will be recognized in the art, the duration of treatment can and will vary depending on the growth and health status of the animal.

[0039] The following numbered examples are contemplated and are non-limiting:

[0040] 1. A composition for improving the utilization of indigestible fiber (NDF) in the feed of a non-human animal and reducing the anti-nutritional effects of the NDF, the composition comprising a basal animal diet supplemented with lysozyme.

[0041] 2. The composition of clause 1, any other suitable clause, or any combination of suitable clauses, wherein the composition is a feed composition or a feed additive composition.

[0042] 3. The composition of clause 1 or clause 2, any other suitable clause, or any combination of suitable clauses, wherein the lysozyme is not a CH-type lysozyme.

[0043] 4. The composition of any one of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the lysozyme of the present disclosure is not a GH-25 lysozyme.

[0044] 5. The feed composition of any one of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the lysozyme is a C-type lysozyme.

[0045] 6. The feed composition of any one of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the lysozyme is a GH-22 lysozyme.

[0046] 7. The feed composition of any one of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the lysozyme is an egg white lysozyme.

[0047] 8. The feed composition of clause 7, any other suitable clause, or any combination of suitable clauses, wherein the concentration of egg white lysozyme in the feed composition ranges from about 0.1 lb / ton to about 0.5 lb / ton.

[0048] 9. The feed composition of clause 7, any other suitable clause, or any combination of suitable clauses, wherein the concentration of egg white lysozyme in the feed composition ranges from about 0.001% w / w to about 0.5% w / w.

[0049] 10. The feed composition of clause 8 or clause 9, any other suitable clause, or any combination of suitable clauses, wherein the animal is a non-ruminant animal.

[0050] 11. The feed composition of any one of clauses 8 to 10, any other suitable clause, or any combination of suitable clauses, wherein the animal is a pig.

[0051] 12. The feed composition of any one of clauses 8 to 11, any other suitable clause, or any combination of suitable clauses, wherein the animal is a nursery pig.

[0052] 13. A method of increasing NDF utilization and decreasing the anti-nutritional effects of the NDF in a non-human animal, the method comprising orally administering to the animal a composition according to any one of clauses 1 to 12, any other suitable clause, or any combination of suitable clauses.

[0053] 14. The method of clause 13, any other suitable clause, or any combination of suitable clauses, wherein the method increases NDF digestibility by at least 36%.

[0054] 15. The method of clause 13 or clause 14, any other suitable clause, or any combination of suitable clauses, wherein the method reduces undigested NDF of the DDGS by at least 13%.

[0055] 16. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases NDF utilization in the presence of a pathogen or in the absence of a pathogen.

[0056] 17. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof.

[0057] 18. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases ADG, ADFI, feed conversion, increases body weight, decreases mortality, decreases total removal, or any combination thereof.

[0058] 19. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases apparent total digestive tract digestibility (ATTD) of dry matter, increases ATTD of total energy, increases ATTD of nitrogen.

[0059] 20. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method reduces the indigestible carbohydrate fraction in the feed composition.

[0060] 21. A method of increasing growth performance, survivability, or growth performance and survivability of a non-human animal, the method comprising orally administering to the animal a composition according to any of clauses 1 to 12, any other suitable clause, or any combination of suitable clauses.

[0061] 22. The method of clause 21, any other suitable clause, or any combination of suitable clauses, wherein the method increases growth performance, survivability, or growth performance and survivability of the animal regardless of the disease status of the animal.

[0062] 23. The method of clause 21 or clause 22, any other suitable clause, or any combination of suitable clauses, wherein the method increases nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof.

[0063] 24. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases ADG, ADFI, feed conversion, increases weight, decreases mortality, decreases total removal, or any combination thereof.

[0064] 25. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases apparent total digestive tract digestibility of dry matter (ATTD), increases ATTD of total energy, increases ATTD of nitrogen.

[0065] 26. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method increases growth performance, survivability, or growth performance and survivability without affecting the proliferation of a pathogen in the animal.

[0066] 27. The method of any of the preceding clauses, any other suitable clause, or any combination of suitable clauses, wherein the method reduces the indigestible carbohydrate fraction in the feed composition.

[0067] All patents and publications mentioned in this specification are indicative of the levels of those skilled in the art to which the disclosure pertains. All patents and publications are herein incorporated by reference to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference.

[0068] The publications discussed throughout the specification are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present application is not entitled to antedate this disclosure by virtue of prior application.

[0069] Examples

[0070] The following examples are included to demonstrate the present disclosure. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventors to function well in the practice of the disclosure. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made to the techniques without departing from the spirit and scope of the disclosure, and, accordingly, all such changes are contemplated as being within the ambit of the present disclosure, and all such changes are deemed to be within the present disclosure.

[0071] Example 1

[0072] Feeding egg white lysozyme (GH-22) to nursery pigs challenged with pathogenic E. coli did not show a beneficial response Egg white lysozyme improved in vitro fiber digestibility of feed

[0073] A study was conducted to evaluate the effects of feeding egg white lysozyme to weaned piglets under E. coli F18 challenge on growth performance and signs of diarrhea. A total of 36 weaned castrated boars aged 19–21 days and weighing 10 lb–16 lb were randomly assigned to one of three dietary treatments (Table 1). From day 6 to day 11 post-inoculation (dpi), the pigs were fed the experimental diet, with a 6-day acclimatization period and an 11-day post-challenge period. On day 0 and day 1 post-inoculation, each pig was orally inoculated with 5 ml (approximately 2.0 × 10⁻⁶ ml) via a polyethylene tube attached to a syringe placed at the back of the mouth. 8 / mL, of which the total excitation amount is 10 9 Freshly grown E. coli F18 inoculum (CFU). Continue feeding pigs their experimental diet and monitor them for 11 days after challenge.

[0074] Table 1. Experimental Dietary Treatments

[0075]

[0076] This *E. coli* challenge model induced diarrhea in over 75% of pigs fed the control diet and the diet containing egg white lysozyme, with signs of severe diarrhea (Table 2). In this study, feeding the diet with pharmacological levels of Zn resulted in 0% mortality, best growth performance, minimal signs of diarrhea, and the lowest frequency of diarrhea. However, compared to the control and ZnO, feeding with egg white lysozyme did not show any beneficial response to growth performance, survival rate, or diarrhea severity. This lack of response to egg white lysozyme is distinct from previous findings that microbial fermentation of lysozyme (GH-25) improves growth performance in pigs and chickens due to its antimicrobial efficacy (see U.S. Patent No. 10,568,344).

[0077] Table 2. Effects of feeding egg white lysozyme on weaned piglets stimulated with pathogenic Escherichia coli.

[0078]

[0079]

[0080] a，b The means of those without a common superscript are different (P < 0.05).

[0081] x，y Means without a common superscript tend to differ (P < 0.10)

[0082] 1 Initially, BW was used as a covariate in all analyses.

[0083] 2Subjective fecal scores were graded on a 0-3 scale: 0 = no signs of diarrhea; 3 = severe signs of diarrhea.

[0084] Example 2

[0085] Feeding egg white lysozyme improved nutrient digestibility and growth performance without affecting the proliferation of E. coli in nursery pigs

[0086] To determine the effect of egg white lysozyme on feed fiber digestibility, corn, corn DDGS, soybean meal, and swine grower-finisher complete feed were mixed with different concentrations of lysozyme from 0% to 0.25%. Each mixture was prepared in duplicate. Each replicate sample was digested using a standardized in vitro rumen procedure (Sawyer and Goeser, 2013) and analyzed for neutral detergent fiber (NDF) content and digestibility in triplicate at Rock River Laboratory (Watertown, Wisconsin).

[0087] The concentration of neutral detergent fiber within each ingredient was similar regardless of lysozyme concentration (Table 3). Inclusion of lysozyme as low as 0.005% was able to increase NDF digestibility by at least 36% and decrease undigested NDF of DDGS by at least 13%. In the other ingredients, lysozyme inclusion as low as 0.05% increased NDF digestibility and decreased undigested NDF. These results indicate that egg white lysozyme (GH-22) can improve fiber digestion and utilization in feeds containing NDF.

[0088] Table 3. Inclusion of 0% to 0.25% lysozyme increased neutral detergent fiber (NDF) digestibility of DDGS.

[0089]

[0090] *Mean values within each ingredient were different from control (P < 0.10)

[0091] **Mean values within each ingredient were different from control (P < 0.05)

[0092] Example 3

[0093] E. coli proliferation in nursery pigs Ingredients

[0094] To evaluate the effect of feeding egg white lysozyme to nursery pigs on nutrient digestibility and growth performance, a total of 60 weaned male pigs, 19-21 days of age, 101b to 16 lb were randomly assigned to one of three dietary treatments of 0 lb / ton, 0.2 lb / ton, and 0.4 lb / ton lysozyme in feed (Table 4).

[0095] Table 4. Dietary treatments

[0096]

[0097] Within 10 days post-weaning, pigs were fed the respective experimental diets. The basal diet was formulated to contain 1.98% crude fiber and 5.60% NDF (Table 5). Each pig received an oral gavage of E. coli F18 inoculum on day 7 to simulate the enteric challenge typically seen in nursery pigs under commercial production conditions. On day 10, pigs were euthanized and ileal mucosa tissue was collected for total coliform and enterotoxigenic E. coli (ETEC) plate counts.

[0098] Table 5. Composition of basal diet

[0099] lb / ton Calculated nutrient composition Corn 600 14% moisture Crude protein, % 860.26 Dried whey 21.12 Crude fat, % 417.00 Soybean 46% 5.7 Crude fiber, % 350.00 Hamlet HP300 1.98 NDF, % 150.00 Menhaden SS-fish meal 5.60 ME, kcal / lb 106.00 Pork fat 1531.45 Lysine, % 67.00 21% Monocal 1.52 SW SIdig Lys, % 14.00 Salt 1.36 TID Thr / Ly, % 11.00 Limestone 0.65 TID M+C / Ly, % 9.10 Lysine 985 0.57 TID Try / Ly, % 5.47 Methionine 99% DL 0.17 TID Iso / Ly, % 3.49 Threonine 98.5% 0.61 TID Val / Ly, % 2.18 Choline chloride 60% 0.67 Calcium, % 1.00 Pig trace minerals 0.90 Phosphorus, % 1.50 Vitamin premix 0.75 Ca / P ratio, ratio 1.00 Sel-Plex 600 ppm 1.2 SW dig P, % 0.50 Selenium 006% 0.52 Lactose, % 0.50 Total 15.01 Figure 1 2000.00

[0100] In the presence of pathogenic E. coli, dietary supplementation of lysozyme up to 0.4 lb / ton linearly increased apparent total digestive tract digestibility (ATTD) of dry matter (p=0.05; Figure 2 ), total energy (p=0.03; Figure 3 ), and nitrogen Figure 4 ; P=0.01) for nursery pigs. The increase in nutrient digestibility was accompanied by a linear increase in pig ADG (p<0.001; Feeding lysozyme improved growth performance and survival of commercial nursery pigs. Pigs fed from 0-24 days post weaning ), reduction in fecal score (p=0.01), and diarrhea frequency (Table 6). However, bacterial proliferation remained unchanged across treatments as measured by total coliform and ETEC counts in ileal tissue (Table 6), suggesting very limited antimicrobial efficacy of egg white lysozyme. In addition, linear correlation analysis indicated that ATTD of dry matter, total energy, and nitrogen were all positively correlated with pig ADG (P<0.06) and negatively correlated with fecal score (P<0.05), but not significantly correlated with coliform or ETEC counts (Table 7). These results, along with the findings in Examples 1 and 2, suggest that the beneficial effects of feeding egg white lysozyme to nursery pigs are related to improving the digestibility and utilization of nutrients in the diet, particularly NDF, and that this mechanism is independent of antimicrobial efficacy.

[0101] Table 6. Effect of dietary supplementation of lysozyme on diarrhea signs and ETEC colonization in ileal tissue of nursery pigs

[0102]

[0103] a，b Means without a common superscript differ (P<0.05)

[0104] 2 Subjective fecal score was graded on a 0-3 scale: 0 = no diarrhea signs; 3 = severe diarrhea signs.

[0105] Table 7. Linear correlations between apparent total digestive tract digestibility (ATTD) of dry matter (DM), gross energy (GE), and nitrogen (N) and pig ADG, fecal score, and ileal tissue bacterial counts

[0106]

[0107] Example 4

[0108] Figure 5-8 Meta-analysis of pigs

[0109] Seven studies were conducted following the same procedure to evaluate the effect of feeding lysozyme to commercial nursery pigs at 21-24 days post-weaning on growth performance and survivability. In each experiment, weaned pigs (initial BW = 12.0 ± 0.11 b) were housed in pens with 27 pigs per pen and randomly assigned to either a control diet (Control) or a diet containing lysozyme, with inclusion levels ranging from 0.1 lb / ton to 0.4 lb / ton feed. This resulted in a total of 1,944 pigs in 72 pens for the Control treatment and 1,917 pigs in 71 pens for the lysozyme treatment across the 7 studies. Each study included standard-of-use feed-grade medicaments.

[0110] All studies, except Study 5, included pharmacological levels of Zn (3,000 ppm) and Cu (150 ppm). An example of the basal diet for the early nursery phase (Phase 1 and 2) is shown in Table 8. NDF concentrations ranged from 4.43% to 6.43% of the complete diet. Data from the 7 experiments were pooled for meta-analysis using the MIXED procedure of SAS.

[0111] Figure 5 Feeding lysozyme to weaned pigs at inclusion levels of 0.1-0.4 lb / ton feed for pigs with BW ranging from 12-24 lb consistently and repeatably improved pig growth performance. Specifically, in the meta-analysis, pigs fed lysozyme had an average 8.1% (P < 0.05) Figure 6 ) improvement in ADG compared to the Control in each of the 7 studies. Pig ADFI was improved an average of 2.7% (P < 0.05) Figure 7 ) in 6 of the 7 studies with lysozyme treatment compared to the Control. Feed conversion was improved an average of 4.6% (P < 0.05) Figure 8 ) in each of the 7 studies compared to the Control. The greater ADG in pigs fed lysozyme resulted in an average 0.8 lb heavier BW (P < 0.05) Figure 9).

[0112] As growth performance improved, survival also responded to dietary supplementation with lysozyme. Specifically, total removal (which includes mortality and poor performers) was reduced by 1.2 percentage points for pigs fed lysozyme compared to controls ( Figure 10 ). Data on mortality only were recorded in Study 5, Study 6, and Study 7. Each of these three studies demonstrated improved survival in the lysozyme treatments, averaging 2.1 percentage points higher than controls ( Feeding lysozyme improved growth performance and survival of commercial nursery pigs. Pigs fed from 0 to 51 days post weaning ). Meta-analysis of early nursery performance in the seven studies demonstrated that consistent and repeatable feeding of lysozyme improved survival and growth performance of nursery pigs fed diets containing 4% to 6% NDF.

[0113] Table 8. Examples of early nursery diets in studies included in the meta-analysis.

[0114]

[0115]

[0116] Example 5

[0117] Meta-analysis Figure 11

[0118] Three of the seven studies described in Example 4 were continued into the late nursery period to evaluate the effect of feeding lysozyme for 49 to 51 days post-weaning (approximate BW 12 lb to 52 lb) on growth performance and survival of commercial nursery pigs. Examples of the base diets for the late nursery period (Stages 3 and 4) are shown in Table 9. The NDF concentration in the late nursery diets was 9.48% to 10.43%. Data from the three experiments were pooled for meta-analysis using the MIXED procedure of SAS.

[0119] Figure 12 The data in Table 10 demonstrate that feeding nursery pigs feed containing lysozyme at inclusion levels of 0.1-0.4 lb / ton of feed consistently and repeatable improved pig growth performance for pigs with BW of 12-52 lb. Specifically, in the meta-analysis, the finishing nursery ADG of pigs fed lysozyme was improved by an average of 4.9% (P < 0.10) in each of the three studies compared to controls. The finishing nursery feed conversion was improved by an average of 3.1% (P < 0.10) in each of the three studies compared to controls. The greater ADG in pigs fed lysozyme resulted in an average of 1.4 lb heavier finishing nursery BW (P < 0.05) compared to controls.

[0120] With improved growth performance, there were also reductions in nursery mortality, total removals, and the percentage of pigs medically treated in each study in the research ​ This meta-analysis of complete nursery performance in the 3 studies indicates that consistent and repeatable feeding of lysozyme improves survival and growth performance of nursery pigs (BW 12 lb to 52 lb) fed diets containing 4% to 10% NDF.

[0121] Table 9

[0122]

[0123]

[0124]

Claims

1. A composition for increasing utilization of non-digestible fiber (NDF) in a feed of a non-human animal and reducing the anti-nutritional effects of the NDF, the composition comprising a basal animal diet supplemented with lysozyme.

2. The composition of claim 1, wherein the composition is a feed composition or a feed additive composition.

3. The composition of claim 1, wherein the lysozyme is not a CH-type lysozyme.

4. The composition of claim 1, wherein the lysozyme is not a GH-25 lysozyme.

5. The feed composition of claim 1, wherein the lysozyme is a C-type lysozyme.

6. The feed composition of claim 1, wherein the lysozyme is a GH-22 lysozyme.

7. The feed composition of claim 1, wherein the lysozyme is an egg white lysozyme.

8. The feed composition of claim 7, wherein the concentration of egg white lysozyme in the feed composition ranges from about 0.1 lb / ton to about 0.5 lb / ton.

9. The feed composition of claim 7, wherein the concentration of egg white lysozyme in the feed composition ranges from about 0.001% w / w to about 0.5% w / w.

10. The feed composition of claim 8, wherein the animal is a non-ruminant animal.

11. The feed composition of claim 8, wherein the animal is a pig.

12. The feed composition of claim 8, wherein the animal is a nursery pig.

13. A method of increasing NDF utilization and reducing the anti-nutritional effects of the NDF in a non-human animal, the method comprising orally administering to the animal the composition of any one of claims 1-12.

14. The method of claim 13, wherein the method increases NDF digestibility by at least 36%.

15. The method of claim 13, wherein the method reduces undigested NDF of DDGS by at least 13%.

16. The method of claim 13, wherein the method increases NDF utilization in the presence or absence of a pathogen.

17. The method of claim 13, wherein the method increases nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof.

18. The method of claim 13, wherein the method increases ADG, ADFI, feed conversion, increases body weight, decreases mortality, decreases total removal, or any combination thereof.

19. The method of claim 13, wherein the method increases apparent total digestive tract digestibility (ATTD) of dry matter, increases ATTD of total energy, increases ATTD of nitrogen.

20. The method of claim 13, wherein the method reduces non-digestible carbohydrate fraction in a feed composition.

21. A method of increasing growth performance, survivability, or growth performance and survivability of a non-human animal, the method comprising orally administering to the animal the composition of any one of claims 1-12.

22. The method of claim 21, wherein the method improves growth performance, survivability, or growth performance and survivability of the animal, regardless of the disease status of the animal.

23. The method of claim 21, wherein the method improves nutrient digestibility, growth performance of the non-human animal, survivability of the non-human animal, or any combination thereof.

24. The method of claim 21, wherein the method improves ADG, ADFI, feed conversion, increases body weight, decreases mortality, decreases total removal, or any combination thereof.

25. The method of claim 21, wherein the method improves apparent total digestive tract digestibility (ATTD) of dry matter, improves ATTD of total energy, improves ATTD of nitrogen.

26. The method of claim 21, wherein the method improves growth performance, survivability, or growth performance and survivability, without affecting the proliferation of a pathogen in the animal.

27. The method of claim 21, wherein the method reduces the fraction of indigestible carbohydrates in the feed composition.

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