Compositions and methods for improving hen welfare with beta-mannanase and probiotics

Abstract

The present disclosure relates to compositions comprising capsicum, functional mineral compounds, and yeast cell wall. Furthermore, methods of feeding the composition to animals such as bovines are provided to demonstrate beneficial effects on the animals. The compositions and methods described herein can benefit overall animal wellbeing and future productivity, especially in younger animals.

Description

COMPOSITIONSMANNANASE AND PROBIOTICSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit under 35 USC § 119(e) of U.S. Provisional Application Serial No. 63 / 400,098, filed on August 23, 2022, the entire disclosure of which is incorporated herein by reference.HELD OF THE INVENTION

[0002] The present disclosure relates to supplements comprising beta-mannanase comprising beta-mannanase and / or a probiotic. Furthermore, methods of administering the supplements to avians such as chickens are provided to demonstrate beneficial effects on the avians. The compositions and methods described herein can benefit overall animal wellbeing and future productivity, especially in egg-laying chickens.BACKGROUND

[0003] Improvement in animal health in avians is an important consideration for the agriculture and food production industries. In modern commercial operations, supplementation of avian feed can be undertaken in order to improve animal health and welfare.

[0004] For instance, the ability to maintain healthy avians that are able to improve their eating, walking, and consumption behavior is typically beneficial to overall avian wellbeing and future productivity. Furthermore, in commercial avian operations, improving egg laying productivity and the resulting quality of eggs is an important consideration. Thus, there exists a need for new compositions and methods to assist with management of avians in order to promote healthy behaviors and improve egg laying characteristics.

[0005] Probiotics can reduce heat stress and abnormal behavior as well as improve health in broiler chickens. Such responses occur from the regulatory power of probiotics under the micro biota-gut-brain axis. Probiotics also alleviate the stress response along the hypothalamic-pituitary-adrenal axis, reducing plasma or brain levels of corticotropin-releasing hormone, adrenocorticotropic hormone, and corticosterone.

[0006] Enzyme supplementation is another strategy that can benefit the gut health status by reducing the impacts of anti-nutritional components. The use of -mannanase can help nonruminant animals dealing with the non-starch polysaccharides, which can reduce nutrient digestibility. Such components are found in plant cell walls and are present in many ingredients largely used in animal feeding, such as soybeans. Among the main hemicelluloses found in plant cell walls are P-mannans, which can also be found on the surface of microorganisms.Thus, the animal's innate immune system is activated when foods that contain P-mannans are ingested, which responds with the proliferation of monocytes, macrophages, dendritic cells, and increased production of cytokines. Such factors generate an unnecessary energy expenditure and an increase in inflammatory responses. By hydrolyzing the P-mannans, this enzyme can improve the digestibility of mannans, increasing the population of beneficial bacteria, improving immunity, digestion and absorption of nutrients, in addition to limiting the proliferation of potential pathogens in the intestine.SUMMARY

[0007] The present disclosure provides supplements comprising beta-mannanase comprising beta-mannanase and / or a probiotic as well as associated methods. The compositions and methods of the present disclosure surprisingly provide a synergistic effect for improve avian behaviors and egg laying qualities. The present disclosure describes supplements comprising beta-mannanase (e.g., Hemicell HT™) and a probiotic (e.g., Protexin™) that can be added to feed to improve animal health and animal welfare.

[0008] Additional features of the present disclosure will become apparent to those skilled in the art upon consideration of illustrative embodiments exemplifying the best mode of carrying out the disclosure as presently perceived.BRIEF DESCRIPTIONS OF THE DRAWINGS

[0009] The detailed description particularly refers to the accompanying figures in which:

[0010] Figure 1 shows examples of eggs with minor presence of feces.

[0011] Figure 2 shows examples of eggs with major presence of feces.

[0012] Figure 3 shows ultramicroscopy images collected in a single eggshell during the first phase.

[0013] Figure 4 shows ultramicroscopy images collected in a single eggshell during the second phase.

[0014] Figure 5 shows ultramicroscopy images collected in a single eggshell during the third phase.DETAILED DESCRIPTION

[0015] In an illustrative aspect, a method comprising a step of administering one or more supplements to a non-human animal is provided. In an embodiment, the animal is an avian. In an embodiment, the avian is selected from the group consisting of a chicken, a turkey, and aduck. In an embodiment, the avian is a chicken. In an embodiment, the chicken is a hen. In an embodiment, the hen is a laying hen. In an embodiment, the avian is present in a cage system.

[0016] In an embodiment, the supplement comprises Hemicell™. In an embodiment, the supplement comprises Hemicell HT™. In an embodiment, the supplement comprises beta- mannanase, also referred to herein as “0- mannanase”.

[0017] In an embodiment, the beta-mannanase is provided by fermentation of Paenibacillus lentus. In an embodiment, the beta-mannanase is administered at a dose between 100 g / ton and 500 g / ton. In an embodiment, the beta-mannanase is administered at a dose of 100 g / ton. In an embodiment, the beta-mannanase is administered at a dose of 200 g / ton. In an embodiment, the beta-mannanase is administered at a dose of 300 g / ton. In an embodiment, the beta-mannanase is administered at a dose of 400 g / ton. In an embodiment, the beta-mannanase is administered at a dose of 500 g / ton.

[0018] In an embodiment, the supplement comprises Protexin™. In an embodiment, the supplement comprises a probiotic supplement. In an embodiment, the probiotic supplement comprises Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactobacillus plantarum, Lactobacillus rhamnosus, Bifidobacterium bifidum, Enterococcus faecium, and Streptococcus thermophiles. In an embodiment, the supplement comprises a probiotic supplement comprising Enterococcus faecium. In an embodiment, the Enterococcus faecium is National Collection of Industrial, Food and Marine Bacteria (NCIMB) culture number 11181

[0019] In an embodiment, the probiotic supplement is administered at a dose between 10 g / ton and 100 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 10 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 20 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 30 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 40 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 50 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 60 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 70 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 80 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 90 g / ton. In an embodiment, the probiotic supplement is administered at a dose of 100 g / ton.

[0020] In an embodiment, the supplements comprise i) beta-mannanase and ii) a probiotic. In an embodiment, the beta-mannanase and the probiotic provide a synergistic effect in improvement in health of the animal.

[0021] In an embodiment, the supplement is present in a feed for the animal. In an embodiment, the feed is a free choice feed.

[0022] In an embodiment, the administration provides an improvement in health of the animal. In an embodiment, the improvement in health of the animal is an improvement in animal welfare. In an embodiment, the improvement in health of the animal comprises an improvement in eating behavior. In an embodiment, the improvement in health of the animal comprises an improvement in walking behavior. In an embodiment, the improvement in health of the animal comprises an improvement in standing behavior. In an embodiment, the improvement in health of the animal comprises an improvement in sitting behavior.

[0023] In an embodiment, the improvement in health of the animal comprises an improvement in liquid consumption behavior. In an embodiment, the improvement in health of the animal comprises an improvement in feather exploring behavior. In an embodiment, the improvement in health of the animal comprises an improvement in head scratching behavior. In an embodiment, the improvement in health of the animal comprises an improvement in wing flapping behavior. In an embodiment, the improvement in health of the animal comprises an improvement in leg stretching behavior. In an embodiment, the improvement in health of the animal comprises an improvement in stretching behavior. In an embodiment, the improvement in health of the animal comprises an improvement in aggressive pecking behavior. In an embodiment, the improvement in health of the animal comprises an improvement in non- aggressive pecking behavior.

[0024] In an embodiment, the improvement in health of the animal comprises a decrease in lesions on the animal. In an embodiment, the lesions are present on the neck. In an embodiment, the lesions are present on the tail. In an embodiment, the lesions are present on the cloaca. In an embodiment, the lesions are present on the comb.

[0025] In an embodiment, the improvement in health of the animal comprises an improvement in intestinal health in the animal. In an embodiment, the improvement in health of the animal comprises prevention of intestinal dysbiosis in the animal.

[0026] In an embodiment, the improvement in health of the animal comprises a reduction in pro-inflammatory effect in the animal. In an embodiment, the improvement in health of the animal comprises increased release of mannan-oiligosaccharides (MOS) in the animal.

[0027] In an embodiment, the administration provides an improvement in egg quality of one of more eggs laid by the animal. In an embodiment, the improvement in egg quality is indicated by a modification of a quality selected from the group consisting of weight, specific gravity, albumen height, albumen weight, yolk height, yolk length, yolk index, yolk weight, Haugh unit, shell weight, shell breaking strength, albumen pH, yolk pH, yolk color score, yolk lightness, yolk redness, yolk yellowness, chroma, and any combination thereof.

[0028] In an embodiment, the improvement in egg quality is indicated by a modification of weight. In an embodiment, the improvement in egg quality is indicated by a modification of specific gravity. In an embodiment, the improvement in egg quality is indicated by a modification of albumen height. In an embodiment, the improvement in egg quality is indicated by a modification of albumen weight. In an embodiment, the improvement in egg quality is indicated by a modification of yolk height. In an embodiment, the improvement in egg quality is indicated by a modification of yolk length. In an embodiment, the improvement in egg quality is indicated by a modification of yolk index. In an embodiment, the improvement in egg quality is indicated by a modification of yolk weight. In an embodiment, the improvement in egg quality is indicated by a modification of Haugh unit. In an embodiment, the improvement in egg quality is indicated by a modification of shell weight. In an embodiment, the improvement in egg quality is indicated by a modification of shell breaking strength. In an embodiment, the improvement in egg quality is indicated by a modification of albumen pH. In an embodiment, the improvement in egg quality is indicated by a modification of yolk pH. In an embodiment, the improvement in egg quality is indicated by a modification of yolk color score. In an embodiment, the improvement in egg quality is indicated by a modification of yolk lightness. In an embodiment, the improvement in egg quality is indicated by a modification of yolk redness. In an embodiment, the improvement in egg quality is indicated by a modification of yolk yellowness. In an embodiment, the improvement in egg quality is indicated by a modification of chroma. In an embodiment, the modification is an increase in measurement. In an embodiment, the modification is a decrease in measurement.

[0029] In an embodiment, the administration provides an improvement in reactive oxygen species (ROS) in the animal. In an embodiment, the improvement in ROS is in the intestine of the animal. In an embodiment, the improvement in ROS is in the oviduct of the animal.

[0030] In an embodiment, the improvement in ROS is indicated by a modification of thiobarbituric acid reactive substances (TBARS), glutathione S-transferase (GST), protein thios (TSH), and any combination thereof. In an embodiment, the improvement in ROS is indicated by a modification of thiobarbituric acid reactive substances (TBARS). In an embodiment, the improvement in ROS is indicated by a modification of glutathione S-transferase (GST). In an embodiment, the improvement in ROS is indicated by a modification of protein thios (TSH). In an embodiment, the modification is an increase in measurement. In an embodiment, the modification is a decrease in measurement.

[0031] In an embodiment, the administration provides an increase in egg laying rate of the animal. In an embodiment, the administration provides an increase in fresh egg weight foreggs laid by the animal. In an embodiment, the administration provides a reduction in coefficient of variability of egg weight for eggs laid by the animal. In an embodiment, the administration provides an increase in an egg component mass for eggs laid by the animal. In an embodiment, the egg component mass is egg mass. In an embodiment, the egg component mass is yolk mass. In an embodiment, the egg component mass is albumen mass. In an embodiment, the egg component mass is egg shell mass.

[0032] In an embodiment, the administration provides an increase in occurrence of clean eggs laid by the animal. In an embodiment, the increase in occurrence of clean eggs is a reduction in the presence of feces on the eggs.

[0033] In an embodiment, the administration provides a modification in eggshell thickness in eggs laid by the animal. In an embodiment, the modification in eggshell thickness is an increase in eggshell thickness. In an embodiment, the modification in eggshell thickness is a decrease in eggshell thickness.

[0034] In an embodiment, the modification in eggshell thickness is observed during a first phase of about 40 weeks. In an embodiment, the modification in eggshell thickness is observed during a second phase of about 44 weeks. In an embodiment, the modification in eggshell thickness is observed during a third phase of about 48 weeks.

[0035] The following numbered embodiments are contemplated and are non-limiting:1. A method comprising a step of administering one or more supplements to a nonhuman animal.2. The method of clause 1 , any other suitable clause, or any combination of suitable clauses, wherein the animal is an avian.3. The method of clause 2, any other suitable clause, or any combination of suitable clauses, wherein the avian is selected from the group consisting of a chicken, a turkey, and a duck.4. The method of clause 2, any other suitable clause, or any combination of suitable clauses, wherein the avian is a chicken.5. The method of clause 4, any other suitable clause, or any combination of suitable clauses, wherein the chicken is a hen.6. The method of clause 5, any other suitable clause, or any combination of suitable clauses, wherein the hen is a laying hen.7. The method of clause 2, any other suitable clause, or any combination of suitable clauses, wherein the avian is present in a cage system.8. The method of any one of clauses 1 to 7, any other suitable clause, or any combination of suitable clauses, wherein the supplement comprises Hemicell HT™.9. The method of any one of clauses 1 to 8, any other suitable clause, or any combination of suitable clauses, wherein the supplement comprises beta-mannanase.10. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is provided by fermentation of Paenibacillus lentus.11. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose between 100 g / ton and 500 g / ton.12. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose of 100 g / ton.13. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose of 200 g / ton.14. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose of 300 g / ton.15. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose of 400 g / ton.16. The method of clause 9, any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase is administered at a dose of 500 g / ton.17. The method of any one of clauses 1 to 16, any other suitable clause, or any combination of suitable clauses, wherein the supplement comprises Protexin™.18. The method of any one of clauses 1 to 17, any other suitable clause, or any combination of suitable clauses, wherein the supplement comprises a probiotic supplement.19. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement comprises Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactobacillus plantarum, Lactobacillus rhamnosus, Bifidobacterium bifidum, Enterococcus faecium, and Streptococcus thermophiles, or wherein the supplement comprises a probiotic supplement comprising Enterococcus faecium, optionally wherein the Enterococcus faecium is National Collection of Industrial, Food and Marine Bacteria (NCIMB) culture number 11181.20. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose between 10 g / ton and 100 g / ton.21. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 10 g / ton.22. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 20 g / ton.23. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 30 g / ton.24. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 40 g / ton.25. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 50 g / ton.26. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 60 g / ton.27. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 70 g / ton.28. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 80 g / ton.29. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 90 g / ton.30. The method of clause 18, any other suitable clause, or any combination of suitable clauses, wherein the probiotic supplement is administered at a dose of 100 g / ton.31. The method of any one of clauses 1 to 30, any other suitable clause, or any combination of suitable clauses, wherein the supplements comprise i) beta-mannanase and ii) a probiotic.32. The method of clause 31 , any other suitable clause, or any combination of suitable clauses, wherein the beta-mannanase and the probiotic provide a synergistic effect in improvement in health of the animal.33. The method of any one of clauses 1 to 32, any other suitable clause, or any combination of suitable clauses, wherein the supplement is present in a feed for the animal.34. The method of clause 33, any other suitable clause, or any combination of suitable clauses, wherein the feed is a free choice feed.35. The method of any one of clauses 1 to 34, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an improvement in health of the animal.36. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal is an improvement in animal welfare.37. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in eating behavior.38. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in walking behavior.39. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in standing behavior.40. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in sitting behavior.41. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in liquid consumption behavior.42. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in feather exploring behavior.43. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in head scratching behavior.44. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in wing flapping behavior.45. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in leg stretching behavior.46. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in stretching behavior.47. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in aggressive pecking behavior.48. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in non- aggressive pecking behavior.49. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises a decrease in lesions on the animal.50. The method of clause 49, any other suitable clause, or any combination of suitable clauses, wherein the lesions are present on the neck.51. The method of clause 49, any other suitable clause, or any combination of suitable clauses, wherein the lesions are present on the tail.52. The method of clause 49, any other suitable clause, or any combination of suitable clauses, wherein the lesions are present on the cloaca.53. The method of clause 49, any other suitable clause, or any combination of suitable clauses, wherein the lesions are present on the comb.54. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises an improvement in intestinal health in the animal.55. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises prevention of intestinal dysbiosis in the animal.56. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises a reduction in pro- inflammatory effect in the animal.57. The method of clause 35, any other suitable clause, or any combination of suitable clauses, wherein the improvement in health of the animal comprises increased release of mannan- oiligosaccharides (MOS) in the animal.58. The method of any one of clauses 1 to 57, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an improvement in egg quality of one of more eggs laid by the animal.59. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of a quality selected from the group consisting of weight, specific gravity, albumen height, albumen weight, yolk height, yolk length, yolk index, yolk weight, Haugh unit, shell weight, shell breaking strength, albumen pH, yolk pH, yolk color score, yolk lightness, yolk redness, yolk yellowness, chroma, and any combination thereof.60. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of weight.61. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of specific gravity.62. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of albumen height.63. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of albumen weight.64. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk height.65. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk length.66. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk index.67. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk weight.68. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of Haugh unit.69. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of shell weight.70. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of shell breaking strength.71. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of albumen pH.72. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk pH.73. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk color score.74. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk lightness.75. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk redness.76. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of yolk yellowness.77. The method of clause 58, any other suitable clause, or any combination of suitable clauses, wherein the improvement in egg quality is indicated by a modification of chroma.78. The method of any one of clauses 59 to 77, any other suitable clause, or any combination of suitable clauses, wherein the modification is an increase in measurement.79. The method of any one of clauses 59 to 77, any other suitable clause, or any combination of suitable clauses, wherein the modification is a decrease in measurement.80. The method of any one of clauses 1 to 79, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an improvement in reactive oxygen species (ROS) in the animal.81. The method of clause 80, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is in the intestine of the animal.82. The method of clause 80, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is in the oviduct of the animal.83. The method of any one of clauses 80 to 82, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is indicated by a modification of thiobarbituric acid reactive substances (TBARS), glutathione S-transferase (GST), protein thios (TSH), and any combination thereof.84. The method of any one of clauses 80 to 82, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is indicated by a modification of thiobarbituric acid reactive substances (TBARS).85. The method of any one of clauses 80 to 82, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is indicated by a modification of glutathione S-transferase (GST).86. The method of any one of clauses 80 to 82, any other suitable clause, or any combination of suitable clauses, wherein the improvement in ROS is indicated by a modification of protein thios (TSH).87. The method of any one of clauses 83 to 86, any other suitable clause, or any combination of suitable clauses, wherein the modification is an increase in measurement.88. The method of any one of clauses 83 to 86, any other suitable clause, or any combination of suitable clauses, wherein the modification is a decrease in measurement.89. The method of any one of clauses 1 to 88, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an increase in egg laying rate of the animal.90. The method of any one of clauses 1 to 89, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an increase in fresh egg weight for eggs laid by the animal.91. The method of any one of clauses 1 to 90, any other suitable clause, or any combination of suitable clauses, wherein the administration provides a reduction in coefficient of variability of egg weight for eggs laid by the animal.92. The method of any one of clauses 1 to 91, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an increase in an egg component mass for eggs laid by the animal.93. The method of clause 92, any other suitable clause, or any combination of suitable clauses, wherein the egg component mass is egg mass.94. The method of clause 92 or clause 93, any other suitable clause, or any combination of suitable clauses, wherein the egg component mass is yolk mass.95. The method of any one of clauses 92 to 94, any other suitable clause, or any combination of suitable clauses, wherein the egg component mass is albumen mass.96. The method of any one of clauses 92 to 95, any other suitable clause, or any combination of suitable clauses, wherein the egg component mass is egg shell mass.97. The method of any one of clauses 1 to 96, any other suitable clause, or any combination of suitable clauses, wherein the administration provides an increase in occurrence of clean eggs laid by the animal.98. The method of clause 97, any other suitable clause, or any combination of suitable clauses, wherein the increase in occurrence of clean eggs is a reduction in the presence of feces on the eggs.99. The method of any one of clauses 1 to 98, any other suitable clause, or any combination of suitable clauses, wherein the administration provides a modification in eggshell thickness in eggs laid by the animal.100. The method of clause 99, any other suitable clause, or any combination of suitable clauses, wherein the modification in eggshell thickness is an increase in eggshell thickness.101. The method of clause 99, any other suitable clause, or any combination of suitable clauses, wherein the modification in eggshell thickness is a decrease in eggshell thickness.102. The method of any one of clauses 1 to 101, any other suitable clause, or any combination of suitable clauses, wherein the modification in eggshell thickness is observed during a first phase of about 40 weeks.103. The method of any one of clauses 1 to 101, any other suitable clause, or any combination of suitable clauses, wherein the modification in eggshell thickness is observed during a second phase of about 44 weeks.104. The method of any one of clauses 1 to 101, any other suitable clause, or any combination of suitable clauses, wherein the modification in eggshell thickness is observed during a third phase of about 48 weeks.EXAMPLESExample 1 Behavioral Assessments

[0036] For the instant example, birds were evaluated at a commercial farm in Salvador do Sul, state of Rio Grande do Sul, in southern Brazil. The experimental units were randomly selected among hens housed in a commercial farm with about 28 thousand lightweight laying hens (Hyline W 36 lineage, 36 weeks old). The replicates were assigned in a completely randomized design to the four treatments, that included i) control treatment (a basal diet without supplementation with any other additive), ii) ; P-mannanase (control diet supplemented with 300 g / ton of -mannanase), hi) probiotic (control diet supplemented with 50 g / ton of a multi- cepa probiotic additive), and iv) P-mannanase plus probiotic treatment (control diet supplemented with 300 g / ton of P-mannanase and 50 g / ton of a multi-cepa probiotic additive).

[0037] The P-mannanase (Hemicell HT™, Elanco Animal Health) for the instant example was an exogenous enzyme from the fermentation of the Paenibacillus lentus bacteria. The probiotic additive (Protexin™ Concentrate, Elanco Animal Health) for the instant example includes Lactobacillus acidophilus (2.06 x 108UFC / g), Lactobacillus bulgaricus (2.06 x 108UFC / g), Lactobacillus plantarum (1.26 x 108UFC / g), Lactobacillus rhamnosus (2.06 x 108UFC / g), Bifidobacterium bifdum (2.0 x 108UFC / g), Enterococcus faecium (6.46 x 108UFC / g), and Streptococcus thermophilus (4.10 x 108UFC / g).

[0038] The basal diet comprised a com-soybean meal-based feed formulated according to nutritional requirements and inert material (kaolin) was included in basal feed to replace - mannanase and / or probiotic additives. Feed and water were both provided ad libitum throughout the experimental period using nipple drinkers and gutter feeders.

[0039] The birds were housed in conventional sheds, arranged in an east-west direction, with concrete floors and masonry walls complemented with wire mesh to the ceiling. The shed was equipped with side curtains, which were managed according to weather conditions to provide thermal comfort. The average minimum and maximum temperature and air relative humidity values recorded were 18 and 36 °C, and 35.8 and 94.7%, respectively. The lighting regime was composed of 16 hours of light and 8 hours of dark per day.

[0040] The birds remained in galvanized- wire cages (100-cm long x 40-cm wide x 45- cm high, resulting in a floor area of 500 cm2 / hen) throughout the experimental period. Birds were allocated in each cage. Birds were supplemented for 84 days and the assessments were performed in the last week of the trial.

[0041] Behavioral assessments were performed via image capture combined with local feather scoping and comb abnormalities assessments. For the behavior assessment, six birds per treatment (one per cage) were randomly selected for observation. The captured images were carried out for 7 consecutive days, in a period of 15 minutes in the morning (e.g., the highest peak of laying of the birds) and 30 minutes in the afternoon divided into two periods corresponding to the hottest and cooler times of the day. Images were recorded and analyzed by visual counting and frequency methods.

[0042] The lesion scoring was performed through visual scoring attributed to three body regions (neck, tail, and vent) from 25 birds per treatment group that were randomly selected. Possible injuries and different degrees of severity were analyzed on a scale from 0 to 5 (best score being 0, indicating complete plumage and no damage, and the worst score being 5, indicating completely feathered areas with skin lesions). Comb abnormalities were observed in the same birds using a scale from 0 to 3, with the best score being 0 (no evidence of comb abnormalities) and the worst score being 3 (3 or more comb areas with evidence of abnormalities).

[0043] All behavioral tests were carried out in the last week of the experiment, allowing the birds to remain exposed to the treatments for a longer period. The same animal was used only in one of the tests, thus preventing one test from interfering with the result of the other.

[0044] The frequency of the main behaviors observed in laying hens fed P-mannanase and / or probiotics are shown in Table 1.Table 1.Treatments1P-value2. T..rai .t.s C „ont .ro -l HeZmIIi,cell Protexin H .. Avg . . . • .. ,_ (H) _ (P) _ + P time _ Treat Time T x T Eatingtimes were polled together (indicated at ‘Avg treat’. The averages obtained when the fourth treatments were polled together in each observation time are also presented (indicated as ‘Avg time’) and compared within the columnXYZ.2Probability of treatment effect (treat), time of observation (time), and interaction (T x T).

[0045] The times expended (minutes / bird) in each of the main behaviors by laying hens fed P-mannanase and / or probiotics are shown in Table 2.Table 2.Walking^eans followed b different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%. Comparisons were performed among treatments - line A.,B,C,D each observation time and also for averages obtained when the three observation times were polled together (indicated at ‘Avg treat’. The averages obtained when the fourth treatments were polled together in each observation time are also presented (indicated as ‘Avg time’) and compared within the columnXYZ.2Probability of treatment effect (treat), time of observation (time), and interaction (T x T).

[0046] The times expended (minutes / bird) in other behaviors by laying hens fed p- mannanase and / or probiotics are shown in Table 3.Table 3.Avg treat 0.33 0.09 0.01 0.09^eans followed b different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%. Comparisons were performed among treatments - line AB,C,Dwjp]jn each observation time and also for averages obtained when the three observation times were polled together (indicated at ‘Avg treat’. The averages obtained when the fourt treatments were polled together in each observation time are also presented (indicated as ‘Avg time’) and compared within the columnXYZ.2Probability of treatment effect (treat), time of observation (time), and interaction (T x T).

[0047] The frequency (%) of birds with lesions (disregarding the score) observed in groups of laying hens fed P-mannanase and / or probiotics are shown in Table 4.Table 4.

[0048] The lesion score observed in groups of laying hens fed 3-mannanase and / or probiotics are shown in Table 5. Scores from 0 (no lesion) to 5 (severe lesion) were attributed to the birds individually, except for crest in which the scores ranged from 0 (no lesion) to 2(severe lesion). Means (LS / means) followed by different uppercase letters differ statistically at5%.Table 5.Example 2 Egg Quality Assessments

[0049] For the instant example, the egg quality of birds in various treatment groups was evaluated. The egg quality was evaluated during storage periods.

[0050] The experimental units were randomly selected among the hens housed in the commercial farm (about 28,000 light-weight laying hens, 36 weeks old, Hyline W 36 lineage) and assigned to the treatments, that were i) control (basal diet, without supplementation); ii) diet supplemented with 300 g / ton of the P-mannanase Hemicell®; iii) diet supplemented with 50 g / ton of the probiotic additive Protexin®; iv) diet supplemented with 50 g / ton of probiotic additive and 300 g / ton of P-mannanase (Hemicell + Protexin).

[0051] The basal feed was formulated according to the nutritional requirements for the genetic (Hy-Line, 2020). Corn and soybean meal were the main ingredients in the feed formulas. Feed and water were both provided ad libitum throughout the experimental period using nipple drinkers and gutter feeders.

[0052] Three 28-day production cycles were carried out and 480 eggs (120 from each treatment group) were randomly collected on the last day of each cycle for evaluation of egg quality. Cracked eggs were excluded from the experiment.

[0053] Part of the eggs were immediately subjected to quality analysis to represent the characteristics of fresh eggs (zero days of storage). The other eggs were stored at room temperature and randomly separated for quality assessment at each storage interval (7, 14, 21, 28, 35 and 42 days).

[0054] Fifteen eggs from each treatment were randomly chosen and weighed individually at weekly intervals during the storage period. The weight loss (%) of eggs during storage was calculated using the following equation:^

[0055] The albumen height was estimated by the average of three measurements taken at different points on the albumen at a distance of 10 mm from the yolk using a digital caliper (TMX PD - 150, China). Thus, the Haugh Unit (HU) was obtained through the equation proposed by Haugh (1937), where h= thickness of albumen (mm); W= mass of the entire egg (g):

[0056] Yolk width and height (mm) were measured with a digital caliper (TMX PD -150, China) on 15 eggs from each treatment at weekly intervals. After, the yolk index was calculated as:

[0057] Yolk color was determined in fifteen eggs per treatment at weekly intervals using the Roche colorimetric fan (DSM, Sao Paulo, Brazil), with a score ranging from 1 (light yellow) to 15 (reddish orange). Complementarily, a spectrophotometer equipment (Delta Vista model 450G, Delta Color, Sao Leopoldo, Brazil) was also used for this evaluation, which determined colorimetric coordinates of luminosity (L*), red intensity (a*) and yellow intensity (b*). Chroma was estimated considering the following equation:

[0058] After yolk and albumen separation, the dense and the fluid albumen were homogenized for 20 seconds and then the pH was determined using a digital pH meter (Kasvi model k39-2014B, Parana, Brazil) previously calibrated with buffer solutions of pH 4, pH 7 and pH 10. The pH of the yolk was determined using the same pH meter.

[0059] The value of specific gravity was based on Archimedes’ principle, using the equation:

[0060] Total solid content was determined separately in albumen and yolk. Five grams of albumen and yolk were weighed separately in previously dried porcelain crucibles. The albumen and yolk samples were kept in an oven at 60 °C for 12 hours and weighed. After weighing, the samples were kept at 105 °C for 12 hours, and weighed again. Seven eggs from each treatment were evaluated to determine the total solids.

[0061] The shell percentage was obtained after shell separation, washing, drying and weighing. Fifteen eggs from each treatment were evaluated on day 0, 21 and 42 for these variables.

[0062] The substances that react to thiobarbituric acid (TBARS) were used to access the lipid oxidation in a pool of three yolks per treatment collected in three storage periods (0, 21, and 42 days). Only treatments with Hemicell and Protexin isolated were tested due to the previous results obtained in egg quality analysis.

[0063] Table 6 displays a summary of fresh egg quality analyses from laying hens fed P-mannanase and / or probiotics. The values represent averages from the three evaluated periods, considering only fresh eggs.Table 6.Treatments, Hemicell Protexin ,, _ ,, , ,Storage time Control . H + P SE H) (r)1P-value(2Weight 62.74 63.11 62.51 62.71 0.290 0.904Specific gravity (g / ml) 1.006B 1.007A 1.006B 1.006B 0.001 <0.001Albumen height (mm) 8.04 8.06 8.18 8.17 0.104 0.129Albumen weight (g) 36.82 37.39 36.30 36.59 0.239 0.424Yolk height (mm) 17.98B 18.15A 18.27A 18.18A 0.063 0.037Yolk length (mm) 40.67B 41.62A 41.25AB 41.82A 0.118 0.002Yolk index 0.443 0.435 0.443 0.435 0.017 0.184Yolk weight (g) 15.33B 15.70AB 15.45B 16.08A 0.096 0.004Haugh unit 89.40 90.10 89.88 89.55 0.558 0.132Shell weight (g) 5.81B 6.15A 5.96AB 6.11A 0.041 0.09Shell breaking strength (N) 2.47 2.39 2.17 2.25 0.106 0.563Albumen pH 8.41 8.40 8.38 8.44 0.028 0.178Yolk pH 6.04B 5.96A 5.99A 6.00A 0.013 0.002Yolk color score (palette) 5.60B 5.98A 5.77 AB 5.87AB 0.052 0.032Yolk lightness (L* color) 50.86B 50.66B 51.33AB 52.16A 0.161 0.002Yolk redness (a* color) 7.12B 7.66A 7.67A 7.66A 0.100 <0.001Yolk yellowness (b* color) 57.41 58.88 58.93 58.75 0.354 0.122Chroma 57.85B 59.67A 59.67A 59.25A 0.357 0.003Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at f0%.

[0064] Table 7 shows weight loss (g) of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 7.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0065] Table 8 shows weight loss (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 8(H) (P) value2^Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0066] Table 9 shows weight loss (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 9.Only 3 treatments 0.062 0.300Only Hemicell 0.085 0.152Only Protexin 0.055 0.670Day 21 1.523 1.562 1.436 1.719All treatments AB AB A B 0.051 0.022Only 3 treatments 0.062 0.424Only Hemicell 0.058 0.708Only Protexin 0.061 0.402Day 28 2.137 2.300 2.018 2.243All treatments 0.053 0.169Only 3 treatments 0.065 0.170Only Hemicell 0.074 0.241Only Protexin 0.080 0.476Day 35 2.439 2.121 2.539 2.592All treatments 0.052 0.182Only 3 treatments 0.058 0.218Only Hemicell 0.077 0.291Only Protexin 0.057 0.358Day 42 3.675 3.263 3.132 3.382All treatments B B A AB 0.049 0.007Only 3 treatments B A A 0.057 0.003Only Hemicell B A 0.069 0.020Only Protexin B A 0.076 <0.001Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0067] Table 10 shows specific gravity (g / ml) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 10.Treatments.. . Hemicell Protexin „ „ P-Storage time Control(H) (p)H + P SE1Day l 1.066 1.059 1.064 1.057All treatments A B B B 0.007 <0.001Only 3 treatments A B A 0.007 <0.001Only Hemicell A B 0.009 <0.001Only Protexin 0.008 0372Day 7 1.051 1.057 1.041 1.054All treatments 0.259 0.997Only 3 treatments 0.345 0.983Only Hemicell 0.520 0.959Only Protexin 0.503 0.922Day 14 1.054 1.048 1.050 1.049All treatments 0.016 0.653Only 3 treatments 0.016 0.348Only Hemicell 0.025 0.242Only Protexin 0.025 0.381Day 21 1.062 1.050 1.051 1.060All treatments 0.028 0.293Only 3 treatments 0.036 0.345Only Hemicell 0.008 0.226Only Protexin 0.008 0.353Day 28 1.042 1.042 1.039 1.041All treatments 0.018 0.954Only 3 treatments 0.021 0.845Only Hemicell 0.022 0.976Only Protexin 0.026 0.522Day 35 1.042 1.042 1.055 1.057All treatments 0.018 0.954Only 3 treatments 0.021 0.845Only Hemicell 0.022 0.976Only Protexin 0.026 0.622Day 42 1.047 1.056 1.055 1.057All treatments B A A A 0.006 <0.001Only 3 treatments B A A 0.007 <0.001Only Hemicell B A 0.009 <0.001Only Protexin B A 0.009 <0.001Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0068] Table 11 shows specific gravity (g / ml) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 11.Treatments.. . i Hemicell Protexin ,, .. P-Storage time Control H + P SE1yalue2Day l 1.075 1.083 1.073 1.074All treatments B A B B 0.008 <0.001Only 3 treatments B A B 0.010 <0.001Only Hemicell B A 0.011 <0.001Only Protexin 0.010 0.246Day ? 1.069 1.066 1.071 1.066All treatments AB AB A B 0.007 0.026Only 3 treatments AB B A 0.008 0.031Only Hemicell a b 0.008 0.065Only Protexin 0.010 0.415Day 14 1.059 1.059 1.053 1.062All treatments A A B A 0.0065 <0.001Only 3 treatments A A B 0.0071 <0.001Only Hemicell 0.0067 0.526Day 28 1.059 1.063 1.061 1.071All treatments B B B A 0.007 <0.001Only 3 treatments B A AB 0.006 0.040Only Hemicell B A 0.007 0.011Only Protexin 0.007 0.251Day 35 1.036 1.039 1.038 1.038All treatments B A AB AB 0.004 0.022Only 3 treatments B A A 0 0.004 0.016Only Hemicell B A 0.006 0.020Only Protexin B A 0.005 0.011Day 42 1.048 1.049 1.050 1.049All treatments 0.004 0.301Only 3 treatments 0.005 0.143Only Hemicell 0.006 0.657Only Protexin b a 0.006 0.054Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0069] Table 12 shows shows specific gravity (g / ml) of eggs from laying hens fedbeta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 12.Treatments, Hemicell Protexin _T_ P-Storag se time Control H + P SB (H) (P)1, value72Day l 1.059 1.065 1.063 1.160A B A A B 0.004 <0.001Only 3 treatments B A A 0.005 <0.001Only Hemicell B A 0.007 <0.001Only Protexin B A 0.006 0.001Day 7 1.064 1.061 1.068 1.064All treatments BC C A B 0.004 <0.001Only 3 treatments B C A 0.005 <0.001Only Hemicell A B 0.004 0.001Only Protexin B A 0.005 <0.001Day 14 1.052 1.053 1.053 1.056All treatments B AB AB A 0.004 0.036Only 3 treatments 0.005 0.607Only Hemicell 0.006 0.347Only Protexin 0.007 0.742Day 21 1.038 1.046 1.042 1.047All treatments C A B A 0.005 <0.001Only 3 treatments C A B 0.005 <0.001Only Hemicell B A 0.008 <0.001Only Protexin B A 0.005 <0.001Day 28 1.031 1.033 1.031 1.032All treatments 0.003 0.223Only 3 treatments 0.004 0.104Only Hemicell b a 0.005 0.079Only Protexin 0.004 0.780Day 35 1.038 1038 1.040 1.039All treatments 0.004 0.954Only 3 treatments 0.004 0.319Only Hemicell 0.004 0.862Only Protexin 0.005 0.240Day 42 1.032 1.037 1.035 1.037All treatments B A A A 0.005 <0.001Only 3 treatments B A A 0.005 <0.001Only Hemicell B A 0.007 <0.001Only Protexin B A 0.005 0.0031Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0070] Table 13 shows albumen height (mm) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 13.Treatments e. .. , Hemicell Protexin ,, „ P-Storag6e time Control (H) (P) H + P SE1va .lue22Day l 9.34 9.69 9.73 8.55All treatments AB A A B 0.14 0.009Only 3 treatments 0.15 0.524Only Hemicell 0.18 0.347Only Protexin 0.17 0.250Day 7 6.03 5.90 6.33 5.18All treatments A A A B 0.08 <0.001Only 3 treatments 0.08 0.102Only Hemicell 0.08 0.461Only Protexin 0.10 0.161Day 14 4.74 4.30 4.79 4.39All treatments AB B A AB 0.07 0.035Only 3 treatments AB B A 0.09 0.048Only Hemicell B A 0.11 0.044Only Protexin 0.11 0.827Day 21 3.97 3.67 3.86 3.56All treatments A AB AB B 0.05 0.041Only 3 treatments 0.06 0.186Only Hemicell a b 0.08 0.096Only Protexin 0.06 0.433Day 28 3.24 3.16 3.31 3.09All treatments 0.04 0.397Only 3 treatments 0.05 0.511Only Hemicell 0.06 0.538Only Protexin 0.07 0.666Day 35 2.98 2.88 2.77 2.77All treatments 0.04 0.343Only 3 treatments 0.05 0.339Only Hemicell 0.08 0.534Only Protexin 0.06 0.144Day 42 2.70 2.76 2.58 2.71All treatments 0.04 0.545Only 3 treatments 0.04 0.285Only Hemicell 0.05 0.644Only Protexin 0.05 0.298Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0071] Table 14 shows albumen height (mm) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 14.Treatments e. .. , Hemicell Protexin ,, „ P-St r tim C ntr l H + P SE1Day l 6.83 6.65 7.05 6.52All treatments 0.11 0.384Only 3 treatments 0.14 0.519Only Hemicell 0.13 0.504Only Protexin 0.20 0.595Day 7 5.33 4.91 5.26 5.08All treatments A B AB B 0.06 0.064Only 3 treatments A B AB 0.06 0.006Only Hemicell A B 0.08 0.007Only Protexin 0.08 0.710Day 14 3.92 3.58 3.73 3.34All treatments A AB AB B 0.06 0.006Only 3 treatments 0.07 0.163Only Hemicell a b 0.08 0.058Only Protexin 0.09 0.336Day 21 3.23 2.89 3.06 2.75All treatments A AB AB B 0.05 0.007Only 3 treatments A B AB 0.05 0.061Only Hemicell A B 0.07 0.016Only Protexin 0.06 0.219Day 28 2.74 2.58 2.72 2.75All treatments 0.04 0.470Only 3 treatments 0.05 0.406Only Hemicell 0.06 0.242Only Protexin 0.06 0.893Day 35 2.75 2.57 2.71 3.06All treatments AB B B A 0.05 0.004Only 3 treatments 0.04 0.325Only Hemicell 0.06 0.173Only Protexin 0.06 0.762Day 42 2.62 2.67 2.74 2.51All treatments AB AB A B 0.03 0.069Only 3 treatments 0.03 0.404Only Hemicell 0.0o4 0.531Only Protexin 0.04 0.1601Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0072] Table 15 shows albumen height (mm) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 15.Treatments.. „ . , Hemicell Protexin .. P-Storag&e time Control ,T„ H + P SE ) (P)1. (H value22Day l 7.97 7.83 7.56 7.64All treatments 0.07 0.209Only 3 treatments 0.09 0.201Only Hemicell 0.12 0.576Only Protexin 0.11 0.119Day 7 5.49 5.28 5.50 5.25All treatments 0.06 0.303Only 3 treatments 0.07 0.394Only Hemicell 0.09 0.249Only Protexin 0.10 0.975Day 14 4.74 4.30 4.79 4.39All treatments A AB A B 0.05 0.011Only 3 treatments a b a 0.07 0.085Only Hemicell a b 0.08 0.051Only Protexin 0.08 0.925Day 21 3.67 3.13 3.51 3.31All treatments A B AB AB 0.06 0.033Only 3 treatments A B AB 0.07 0.012Only Hemicell A B 0.01 0.016Only Protexin 0.06 0.214Day 28 2.96 2.82 2.93 2.73All treatments a a a b 0.03 0.098Only 3 treatments 0.04 0.424Only Hemicell 0.05 0.245Only Protexin 0.05 0.791Day 35 2.66 2.48 2.52 2.77All treatments AB B B A 0.03 0.018Only 3 treatments 0.04 0.303Only Hemicell a b 0.04 0.057Only Protexin 0.06 0.295Day 42 2.88 2.47 2.48 2.36All treatments A AB AB B 0.04 0.050Only 3 treatments 0.06 0.176Only Hemicell 0.06 0.106Only Protexin 0.08 0.130Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0073] Table 16 shows yolk length (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 16.Treatments, Hemicell Protexin ....| P-Storage time Control H + P SE1value2Day l 40.63 41.25 41.32 41.50All treatments 0.25 0.565Only 3 treatments 0.24 0.467Only Hemicell 0.30 0.319Only Protexin 0.29 0.261Day 7 43.49 44.38 43.50 44.38All treatments 0.23 0.319Only 3 treatments 0.27 0.323Only Hemicell 0.39 0.262Only Protexin 0.27 0.986Day 14 45.02 45.58 44.48 44.52All treatments 0.23 0.285Only 3 treatments 0.24 0.188Only Hemicell 0.31 0.391Only Protexin 0.27 0.328Day 21 49.84 46.85 45.71 47.91All treatments A B B AB 0.42 0.00Only 3 treatments A B B 0.52 0.001Only Hemicell A B 0.73 0.005Only Protexin A B 0.73 0.005Day 28 47.91 48.00 46.71 46.00Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0074] Table 17 shows yolk length (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 17.Only Protexin A B 0.50 0.003Day 35 47.90 48.50 48.47 47.96All treatments 0.28 0.816Only 3 treatments 0.34 0.733Only Hemicell 0.35 0.416Only Protexin 0.45 0.536Day 42 51.59 51.91 51.02 49.76All treatments 0.39 0.238Only 3 treatments 0.48 0.739Only Hemicell 0.56 0.782Only Protexin 0.66 0.680Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0075] Table 18 shows yolk length (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 18.Treatments o. , Hemicell Protexin ,, „ P-Storage time Control H + P SE1yalue2Day l 39.94 41.10 41.40 41.96All treatments B AB AB A 0.19 0.002Only 3 treatments b a a 0.22 0.051Only Hemicell B A 0.26 0.024Only Protexin B A 0.27 0.032Day 7 42.03 42.63 42.76 44.01All treatments B AB AB A 0.23 0.017Only 3 treatments 0.23 0.400Only Hemicell 0.28 0.301Only Protexin 0.28 0.208Day 14 46.63 45.40 45.93 46.21All treatments 0.24 0.355Only 3 treatments 0.31 0.248Only Hemicell a b 0.37 0.099Only Protexin 0.39 0.378Day 21 46.08 46.10 46.18 47.66All treatments 0.25 0.107Only 3 treatments 0.27 0.989Only Hemicell 0.37 0.977Only Protexin 0.29 0.872Day 28 46.08 45.47 47.16 48.99All treatments B B AB A 0.31 <0.001Only 3 treatments 0.33 0.108Only Hemicell 0.30 0.328Only Protexin 0.41 0.217Day 35 48.06 47.96 48.39 47.44All treatments 0.35 0.809Only 3 treatments 0.42 0.913Only Hemicell 0.54 0.928Only Protexin 0.60 0.790Day 42 50.00 49.04 48.80 49.71All treatments 0.35 0.611Only 3 treatments 0.43 0.502Only Hemicell 0.58 0.418Only Protexin _ 0.47 _ 0.206Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0076] Table 19 shows yolk height (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 19.Treatments.. , Hemicell Protexin ,, „ P-Storag6e time Control H + P SE (H) (P)1. value22Day l 18.66 18.44 19.13 18.34All treatments AB AB A B 0.10 0.035Only 3 treatments b b a 0.12 0.060Only Hemicell 0.12 0.367Only Protexin 0.16 0.147Day 7 15.77 15.22 15.36 14.98All treatments A AB AB B 0.10 0.57Only 3 treatments 0.11 0.154Only Hemicell 0.13 0.132Only Protexin 0.15 0.184Day 14 13.15 13.09 13.10 13.05All treatments 0.10 0.987Only 3 treatments 0.12 0.974Only Hemicell 0.14 0.820Only Protexin 0.18 0.873Day 21 11.82 12.08 11.94 11.96All treatments 0.07 0.717Only 3 treatments 0.08 0.520Only Hemicell 0.11 0.295Only Protexin 0.11 0.633Day 28 10.51 10.48 11.21 11.40All treatments B B A A 0.09 <0.001Only 3 treatments B B A 0.10 0.003Only Hemicell 0.11 0.875Only Protexin B A 0.12 0.002Day 35 10.07 10.03 11.01 9.81All treatments B B A B 0.11 <0.001Only 3 treatments B B A 0.13 0.001Only Hemicell 0.14 0.894Only Protexin B A 0.14 <0.001Day 42 9.96 9.51 10.07 10.26All treatments b b a ab 0.11 0.088Only 3 treatments 0.13 0.193Only Hemicell 0.14 0.119Only Protexin 0.17 0.776^Standard error.2Prohability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0077] Table 20 shows yolk height (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 20.TreatmentsDay l 17.88 17.79 17.94 18.11All treatments 0.08 0.273Only 3 treatments 0.09 0.551Only Hemicell 0.11 0.633Only Protexin 0.10 0.215Day 7 15.50 15.77 16.27 16.01All treatments B AB A AB 0.09 0.018Only 3 treatments B AB A 0.11 0.020Only Hemicell 0.13 0.327Only Protexin B A 0.15 0.013Day 14 13.47 13.42 13.78 13.10All treatments 0.10 0.122Only 3 treatments 0.11 0.400Only Hemicell 0.13 0.880Only Protexin 0.15 0.306Day 21 12.33 11.83 12.23 12.17All treatments 0.08 0.180Only 3 treatments a b a 0.09 0.067Only Hemicell 0.13 0.127Only Protexin 0.09 0.510Day 28 10.86 10.46 10.83 10.88All treatments 0.09 0.431Only 3 treatments 0.09 0.205Only Hemicell 0.11 0.103Only Protexin 0.11 0.900Day 35 10.88 11.00 11.39 10.96All treatments 0.09 0.266Only 3 treatments 0.12 0.206Only Hemicell 0.15 0.700Only Protexin A B 0.12 0.040Day 42 9.64 9.59 10.49 10.07All treatments B B A AB 0.10 0.002Only 3 treatments B B A 0.12 0.001Only Hemicell 0.14 0.860Only Protexin _ B _ A _ 0.16 _ 0.007Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0078] Table 21 shows yolk height (mm) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 21.Treatments o. .. „ . , Hemicell Protexin ,, „ P-Stora e time Control H + P SE1Day l 17.59 17.92 17.75 18.11All treatments 0.10 0.333Only 3 treatments 0.12 0.542Only Hemicell 0.15 0.304Only Protexin 0.16 0.636Day 7 15.97 15.70 15.81 16.27All treatments 0.09 0. 194Only 3 treatments 0.10 0.592Only Hemicell 0.13 0.324Only Protexin 0.13 0.564Day 14 14.05 13.61 13.65 13.81All treatments 0.08 0.247Only 3 treatments 0.09 0.131Only Hemicell 0.12 0.102Only Protexin 0.12 0.103Day 21 13.07 12.96 12.67 12.87All treatments 0.10 0.569Only 3 treatments 0.12 0.392Only Hemicell 0.12 0.756Only Protexin 0.13 0.131Day 28 12.39 12.28 12.38 11.95All treatments 0.08 0.221Only 3 treatments 0.08 0.863Only Hemicell 0.08 0.525Only Protexin 0.12 0.961Day 35 11.48 11.28 11.23 11.36All treatments 0.08 0.802Only 3 treatments 0.09 0.557Only Hemicell 0.10 0.373Only Protexin 0.11 0.315Day 42 10.74 10.41 10.79 10.39All treatments 0.10 0.385Only 3 treatments 0.12 0.432Only Hemicell 0.17 0.351Only Protexin 0.14 0.872Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0079] Table 22 shows yolk index of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 22.Standard error. Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0080] Table 23 shows yolk index of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 23.Treatments. - Hemicell ProtexinrT„ P-Storage time Control ( / THT). (ZTPV)lH + P SE1va ,lue22Day l 0.429 0.418 0.433 0.435All treatments ab a b b 0.002 0.083Only 3 treatments 0.003 0.142Only Hemicell 0.003 0.113Only Protexin 0.003 0.600Day 7 0.347 0.369 0.383 0.369All treatments B A A A 0.002 <0.001Only 3 treatments B A A 0.003 <0.001Only Hemicell B A 0.003 0.002Only Protexin B A 0.004 <0.001Day 14 0.293 0.294 0.312 0.289All treatments B B A B 0.002 0.002Only 3 treatments B B A 0.003 0.012Only Hemicell 0.002 0.947Only Protexin B A 0.004 0.020Day 21 0.260819 0.248324 0.270605 0.258164All treatments AB B B AB 0.002 0.032Only 3 treatments AB B B 0.003 0.013Only Hemicell 0.003 0.101Only Protexin 0.003 0.190Day 28 0.214 0.218 0.228 0.225All treatments 0.002 0.286Only 3 treatments 0.003 0.156Only Hemicell 0.003 0.602Only Protexin 0.003 0.074Day 35 0.228 0.232 0.236 0.229All treatments 0.002 0.730Only 3 treatments 0.003 0.631Only Hemicell 0.004 0.622Only Protexin 0.003 0.240Day 42 0.186 0.185 0.205 0.202All treatments B B A A 0.003 0.023Only 3 treatments AB B A 0.003 0.033Only Hemicell 0.003 0.907Only Protexin b a 0.005 0.064Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0081] Table 24 shows yolk index of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 24.Treatments, Hemicell Protexin .. , P-Storag6e time Control (H) (P) H + P SE1va .lue22Day l 0.441 0.436 0.432 0.432All treatments 0.002 0.623Only 3 treatments 0.003 0.571Only Hemicell 0.004 0.618Only Protexin 0.004 0.340Day 7 0.380 0.369 0.366 0.370All treatments 0.003 0.407Only 3 treatments 0.003 0.266Only Hemicell 0.003 0.148Only Protexin 0.004 0.155Day 14 0.314 0.299 0.301 0.299All treatments 0.002 0.156Only 3 treatments 0.003 0.146Only Hemicell 0.004 0.108Only Protexin 0.004 0.147Day 21 0.284 0.271 0.275 0.270All treatments 0.003 0.486Only 3 treatments 0.004 0.435Only Hemicell 0.003 0.291Only Protexin 0.003 0.197Day 28 0.271 0.271 0.263 0.243All treatments A A AB B 0.003 0.003Only 3 treatments 0.003 0.589Only Hemicell 0.002 0.947Only Protexin 0.004 0.399Day 35 0.239 0.235 0.229 0.240All treatments 0.002 0.434Only 3 treatments 0.003 0.416Only Hemicell 0.003 0.598Only Protexin 0.004 0.27Day 42 0.220 0.214 0.222 0.210All treatments 0.003 0.483Only 3 treatments 0.003 0.682Only Hemicell 0.005 0.575Only Protexin 0.003 0.848Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0082] Table 25 shows yolk weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 25.yStandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0083] Table 26 shows yolk weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 26.' Slandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0084] Table 27 shows yolk weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 27.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0085] Table 28 shows Haugh unit of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 28.Only Hemicell b a 1.06 0.080Only Protexin 0.82 0.921Day 7 76.76 75.55 78.39 70.26All treatments A A A B 0.74 <0.001Only 3 treatments 0.73 0.292Only Hemicell 0.84 0.484Only Protexin 0.38 0.369Day 14 85.55 60.84 67.17 61.36All treatments AB B A 0.89 0.126Only 3 treatments 1.08 0.044Only Hemicell 1.44 0.120Only Protexin 1.16 0.496Day 21 58.18 53.70 53.91 51.21All treatments A AB AB B 0.94 0.075Only 3 treatments 1.09 0.176Only Hemicell 1.48 0.132Only Protexin 1.16 0.163Day 28 48.49 44.55 48.51 48.09All treatments 1.02 0.458Only 3 treatments 1.17 0.294Only Hemicell 1.52 0.200Only Protexin 1.49 0.994Day 35 44.22 42.70 42.03 42.41All treatments 1.05 0.309Only 3 treatments 1.24 0.185Only Hemicell 1.56 0.152Only Protexin 1.63 0.112Day 42 39.62 41.97 39.88 42.16All treatments 0.93 0.582Only 3 treatments 109 0.857Only Hemicell 1.56 0.466Only Protexin 1.22 0.117Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0086] Table 29 shows Haugh unit of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage Limes in a second trial.Table 29.Treatments.. z . . Hemicell Protexin .. ,, P-Storage time Control H + P SE1Day l 84.42 82.53 85.31 79.85All treatments A AB A B 0.58 0.003Only 3 treatments 0.62 0.181Only Hemicell 0.76 0.221Only Protexin 0.76 0.566Day 7 70.58 69.01 69.00 66.96All treatments 0.69 0.343Only 3 treatments 0.81 0.195Only Hemicell 0.90 0.140Only Protexin 1.00 0.443Day 14 58.24 53.89 57.42 48.30All treatments A AB A B 1.01 0.001Only 3 treatments 1.07 0.213Only Hemicell 1.34 0.106Only Protexin 1.23 0.746Day 21 47.09 43.03 46.60 41.53All treatments A A A B 0.92 0.094Only 3 treatments 1.12 0.287Only Hemicell 1.41 0.154Only Protexin 1.40 0.863Day 28 41.03 39.52 41.68 38.71All treatments 0.77 0.543Only 3 treatments 0.87 0.587Only Hemicell 1.18 0.533Only Protexin 1.04 0.759Day 35 43.40 41.17 39.60 46.44All treatments ABA B B A 0.88 0.030Only 3 treatments 0.96 0.292Only Hemicell 1.12 0.331Only Protexin 1.35 0.163Day 42 38.10 41.18 39.96 38.45All treatments 0.85 0.593Only 3 treatments 1.09 0.557Only Hemicell 1.39 0.280Only Protexin 1.31 0.497Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0087] Table 30 shows Haugh unit of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 30.Treatments. Hemicell Protexin P-Storag6e time Control H + P SE1. (H) (P) value22Day l 86.93 87.22 87.38 87.14All treatments 0.42 0.990Only 3 treatments 0.55 0.58Only Hemicell 0.76 0.858Only Protexin 0.66 0.770Day 7 71.65 68.71 70.85 67.55All treatments 0.88 0.134Only 3 treatments 0.78 0.297Only Hemicell 0.98 0.139Only Protexin 0 96 0 5771Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0088] Table 31 shows albumen weight (g) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 31.Only Protexin 0.55 0.200Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0089] Table 32 shows albumen weight (g) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 32.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0090] Table 33 shows albumen weight (g) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 33.Only Hemicell 0.62 0.153Only Protexin 0.61 0.966Day 21 38.56 38.43 36.77 37.07All treatments 0.43 0.342Only 3 treatments 0.52 0.294Only Hemicell 0.66 0.926Only Protexin 0.58 0.132Day 28 36.07 37.92 37.18 35.89All treatments 0.46 0.369Only 3 treatments 0.56 0.411Only Hemicell 065 0.162Only Protexin 0.71 0.445Day 35 34.49 35.36 34.86 34.55All treatments 0.43 0.900Only 3 treatments 0.49 0.92Only Hemicell 0.68 0.536Only Protexin 0.61 0.775Day 42 33.99 33.36 33.12 33.80All treatments 0.43 0.891Only 3 treatments 0.49 0.761Only Hemicell 0.57 0.589Only Protexin 0.65 0.518Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0091] Table 34 shows shell weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 34.Treatments... .. , Hemicell Protexin ,, „ P-Storage time Control H + P SE1yalue2Day l 5.78 6.07 6.05 6.03All treatments 0.07 0.482Only 3 treatments 0.09 0.371Only Hemicell 0.12 0.262Only Protexin 0.07 0.071Day 7 6.17 6.10 6.15 6.19All treatments 0.04 0.919Only 3 treatments 0.05 0.873Only Hemicell 0.07 0.635Only Protexin 0.06 0.901Day 14 6.01 6.02 6.18 6.07All treatments 0.05 0.636Only 3 treatments 0.06 0.489Only Hemicell 0.06 0.931Only Protexin 0.08 0.300Day 21 5.81 6.08 5.93 6.11yStandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0092] Table 35 shows shell weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 35.Day 35 5.95 5.90 5.75 6.13All treatments 0.05 0.112Only 3 treatments 0.05 0.278Only Hemicell 0.06 0.737Only Protexin 0.06 0.130Day 42 6.22 6.26 6.01 6.39All treatments 0.06 0.245Only 3 treatments 0.07 0.394Only Hemicell 0.09 0.859Only Protexin 0.10 0.307Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0093] Table 36 shows shell weight (g) of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 36.Treatments.. „ . , Hemicell Protexin .. _ P-Storagse time Control (H) (P) H + P SE1val .ue2,Day l 5.40 6.18 5.90 6.13All treatments B A AB A 0.07 <0.001Only 3 treatments B A A 0.09 0.002Only Hemicell B A 0.12 0.001Only Protexin B A 0.10 0.016Day 7 5.32 6.26 5.84 4.97All treatments C A B AB 0.06 <0.001Only 3 treatments C A B 0.08 <0.001Only Hemicell B A 0.11 <0.001Only Protexin B A 0.09 0.003Day 14 5.42 6.17 6.03 6.09All treatments B A A A 0.07 <0.001Only 3 treatments B A A 0.09 0.001Only Hemicell B A 0.11 0.001Only Protexin B A 0.10 0.002Day 21 5.70 6.03 5.83 6.21All treatments B AB B A 0.05 0.003Only 3 treatments B A B 0.05 0.089Only Hemicell b a 0.08 0.051Only Protexin 0.06 0.320Day 28 5.76 6.18 6.09 6.17All treatments 0.06 0.101Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0094] Table 37 shows shell breaking strength (N) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 37.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0095] Table 38 shows shell breaking strength (N) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 38.yStandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0096] Table 39 shows shell breaking strength (N) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 39.' Slandard error. Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0097] Table 40 shows albumen pH of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 40.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0098] Table 41 shows albumen pH of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 41.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0099] Table 42 shows albumen pH of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 42.Treatments, Hemicell Protexin , P-Storag6e time Control (H) (P) H + P SE1va .lue22Day l 8.13 8.05 8.09 8.10All treatments 0.01 0.178Only 3 treatments 0.01 0.163Only Hemicell 0.02 0.101Only Protexin 0.02 0.481Day 7 8.30 8.35 8.35 8.37All treatments 0.01 0.127Only 3 treatments 0.01 0.239Only Hemicell 0.01 0.171Only Protexin 0.01 0.155Day 14 8.46 8.48 8.49 8.50All treatments 0.01 0.175Only 3 treatments 0.01 0.552Only Hemicell 0.01 0.545Only Protexin 0.01 0.361Day 21 8.57 8.59 8.60 8.58All treatments 0.01 0.417Only 3 treatments 0.01 0.274Only Hemicell 0.01 0.437Only Protexin 0.01 0,125Day 28All treatments 0.01Only 3 treatments 0.01Only Hemicell 0.01Only Protexin 0.01Day 35 8.75 8.77 8.78 8.76All treatments 0.01 0.490Only 3 treatments 0.01 0.336Only Hemicell 0.01 0.403Only Protexin 0.01 0.192Day 42 9.08 9.09 9.09 9.09All treatments 0.01 0.946Only 3 treatments 0.01 0.879Only Hemicell 0.01 0.669Only Protexin 0.01 0.704Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0100] Table 43 shows yolk pH of eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 43.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0101] Table 44 shows yolk pH of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 44.y' Slandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0102] Table 45 shows yolk pH of eggs from laying hens fed beta-mannanase(Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 45.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0103] Table 46 shows yolk color score (palette) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 46.All treatments 0.10 0.100Only 3 treatments 0.13 0.254Only Hemicell b a 0.13 0.075Only Protexin 0.17 0.999Day 7 4.87 4.87 4.80 5.21All treatments 0.07 0.221Only 3 treatments 0.08 0.935Only Hemicell 0.10 0.999Only Protexin 0.10 0.764Day 14 5.42 5.13 6.00 5.27All treatments B B A B 0.08 <0.001Only 3 treatments B B A 0.10 0.001Only Hemicell 0.08 0.102Only Protexin B A 0.13 0.031Day 21 6.36 6.07 6.20 6.07All treatments 0.08 0.574Only 3 treatments 0.08 0.430Only Hemicell 0.10 0.166Only Protexin 0.12 0.525Day 28 5.53 6.64 6.60 6.62All treatments B A A A 0.07 0.001Only 3 treatments B A A 0.08 <0.001Only Hemicell B A 0.11 <0.001Only Protexin B A 0.10 0.001Day 35 6.38 6.67 6.64 6.29All treatments 0.08 0.289Only 3 treatments 0.09 0.446Only Hemicell 0.10 0.202Only Protexin 0.13 0.348Day 42 6.18 6.91 6.82 6.38All treatments B A A AB 0.08 <0.001Only 3 treatments B A A 0.09 0.001Only Hemicell B A 0.12 0.002Only Protexin B A 0.10 0.001Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0104] Table 47 shows yolk color score (palette) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 47.Treatments... , Hemicell Protexin .. „ P-S to rag6e time Control , (T_ H + P SE H) (P)1, value72Day l 5.33 6.00 5.93 5.60All treatments B A A AB 0.07 0.004Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0105] Table 48 shows yolk color score (palette) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 48.Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0106] Table 49 shows yolk lightness (L* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 49.Only Protexin 0.45 0.849Day 7 58.18 58.72 57.51 57.20All treatments a a ab b 0.21 0.051Only 3 treatments 0.24 0.125Only Hemicell 0.27 0.336Only Protexin 0.32 0.300Day 14 57.60 58.70 58.27 61.36All treatments 0.61 0.133Only 3 treatments 0.61 0.772Only Hemicell 0.89 0.548Only Protexin 0.39 0.408Day 21 57.13 58.45 57.48 58.16All treatments B B AB AB 0.18 0.036Only 3 treatments B A AB 0.21 0.032Only Hemicell B A 0.23 0.003Only Protexin 0.26 0.512Day 28 55.27 58.72 57.86 58.54All treatments B A A A 0.313 <0.001Only 3 treatments B A A 0.377 <0.001Only Hemicell B A 0.505 <0.001Only Protexin B A 0.491 0.006Day 35 55.38 58.60 57.53 56.25All treatments C A AB BC 0.31 0.001Only 3 treatments B A A 0.35 <0.001Only Hemicell B A 0.49 <0.001Only Protexin B A 0.43 0.010Day 42 57.77 59.14 58.96 58.53All treatments 0.22 0.121Only 3 treatments b a ab 0.26 0.071Only Hemicell B A 0.32 0.029Only Protexin b a 0.35 0.0921Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0107] Table 50 shows yolk lightness (L* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 50.Treatments.. z , . Hemicell Protexin .. ,, P-Storage time Control H + P SE1Day l 50.99 51.65 52.06 53.73All treatments B B AB A 0.27 0.001Only 3 treatments 0.27 0.300Only Hemicell 0.32 0.311Only Protexin a 0.30 0.079Day 7 55.77 56.35 56.45 56.07All treatments 0.23 0.741Only 3 treatments 0.26 0.544Only Hemicell 0.26 0.276Only Protexin 0.35 0.343Day 14 57.87 57.87 58.54 59.04All treatments 0.19 0.101Only 3 treatments 0.22 0.377Only Hemicell 0.29 0.995Only Protexin 0.30 0.284Day 21 56.62 58.01 58.93 58.32All treatments B AB A AB 0.25 0.008Only 3 treatments B AB A 0.32 0.008Only Hemicell b a 0.41 0.099Only Protexin B A 0.43 0.005Day 28 58.25 58.11 58.31 59.82All treatments B B B A 0.18 0.003Only 3 treatments 0.16 0.885Only Hemicell 0.21 0.757Only Protexin 0.18 0.870Day 35 58.73 58.82 58.89 58.97All treatments 0.17 0.970Only 3 treatments 0.21 0.954Only Hemicell 0.28 0.878Only Protexin 0.24 0.750Day 42 59.07 58.84 59.10 59.88All treatments 0.22 0.385Only 3 treatments 0.24 0.899Only Hemicell 0.37 0.762Only Protexin 0.26 0.948Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0108] Table 51 shows yolk lightness (L* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 51.Treatments, Hemiccll Protexin¥¥„ P-Storage time Control H + P SE1Day l 50.26 50.01 50.45 50.88All treatments 0.23 0.777Only 3 treatments 0.25 0.776Only Hemicell 0.29 0.677Only Protexin 0.32 0.768Day 7 53.37 56.34 54.60 55.78All treatments B A AB 0.32 0.003Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0109] Table 52 shows yolk redness (a* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 52.Only Hemicell b a 0.19 0.081Only Protexin 0.18 0.572Day 14 7.21 7.62 7.28 6.69All treatments 0.12 0.067Only 3 treatments 0.14 0.494Only Hemicell 0.17 0.265Only Protexin 0.10 0.860Day 21 7.02 7.41 6.71 6.36All treatments 0.15 0.112Only 3 treatments 0.18 0.298Only Hemicell 0.17 0.266Only Protexin 0.12 0.534Day 28 6.57 7.46 7.15 6.20All treatments AB A A B 0.1 0.003Only 3 treatments B A AB 0.14 0.034Only Hemicell B A 0.17 0.007Only Protexin 0.16 0.201Day 35 5.80 6.79 7.28 6.27All treatments B AB A AB 0.15 0.005Only 3 treatments B AB A 0.18 0.005Only Hemicell B A 0.24 0.040Only Protexin B A 0.17 0.003Day 42 7.01 6.95 7.45 7.24All treatments 0.17 0.757Only 3 treatments 0.13 0.245Only Hemicell 0.16 0.862Only Protexin 0.14 0.203Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0110] Table 53 shows yolk redness (a* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 53.Treatments.. , Hemicell Protexin ,, „ P-Storage time Control H + P SE1Day l 6.98 7.20 7.97 6.02All treatments AB A A B 0.16 <0.001Only 3 treatments B B A 0.13 0.003Only Hemicell 0.14 0.460Only Protexin B A 0.10 0.002Day 7 6.17 6.98 7.16 6.87All treatments B A A AB 0.11 0.017Only 3 treatments B A A 0.14 0.008Only Hemicell B A 0.18 0.025Only Protexin B A 0.12 0.006Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0111] Table 54 shows yolk redness (a* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 54.All treatments 0.14 0.370Only 3 treatments 0.17 0.900Only Hemicell 0.20 0.920Only Protexin 0.17 0.751Day 21 6.09 6.04 6.38 5.09All treatments AB AB A B 0.18 0.058Only 3 treatments 0.21 0.777Only Hemicell 0.28 0.937Only Protexin 0.10 0.512Day 28 5.49 6.05 6.11 5.93All treatments 0.14 0.420Only 3 treatments 0.18 0.332Only Hemicell 0.22 0.233Only Protexin 0.12 0.235Day 35 5.58 6.33 6.22 5.29All treatments AB AB A B 0.14 0.024Only 3 treatments b a a 0.14 0.088Only Hemicell b a 0.20 0.069Only Protexin 0.10 0.125Day 42 5.39 6.38 5.89 5.48All treatments B A AB B 0.12 0.011Only 3 treatments B A AB 0.12 0.002Only Hemicell B A 0.16 0.001Only Protexin b a 0.11 0.097Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0112] Table 55 shows yolk yellowness (b* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a first trial.Table 55.Treatments.. , , . , Hemicell Protexin .. P-Storag6e time Control ,T„ H + P SE (H) (P)1, , value2Day l 6.05 6.16 5.96 6.12All treatments 0.33 0.167Only 3 treatments AB A B 0.34 0.060Only Hemicell 0.43 0.224Only Protexin 0.33 0.178Day 7 5.98 6.12 6.00 5.98All treatments 0.35 0.445Only 3 treatments 0.40 0.302Only Hemicell 0.45 0.118Only Protexin 0.53 0.880Day 14 6.02 6.40 5.98 5.98All treatments B A B B 0.53 0.013Only 3 treatments B A B 0.67 0.018Only Hemicell B A 0.89 0.030Only Protexin 0.64 0.799Day 21 5.96 6.23 5.96 5.85All treatments AB A AB B 0.43 0.011Only 3 treatments B A B 0.48 0.025Only Hemicell B A 0.50 0.005Only Protexin 0.55 0.997Day 28 5.63 6.16 5.98 5.87All treatments C A AB BC 0.45 <0.001Only 3 treatments B A A 0.55 <0.001Only Hemicell B A 0.78 <0.001Only Protexin B A 0.85 0.006Day 35 5.53 5.83 5.61 5.36All treatments ab a ab b 0.66 0.072Only 3 treatments 0.73 0.204Only Hemicell b a 0.90 0.089Only Protexin 0.97 0.688Day 42 5.55 5.67 5.77 5.69All treatments 0.60 0.666Only 3 treatments 0.72 0.487Only Hemicell 0.88 0.507Only Protexin 0.92 0.252Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0113] Table 56 shows yolk yellowness (b* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a second trial.Table 56.Treatments o. , Hemicell Protexin ,, „ ,1T,. P-Storage time Control H + P SE1Day l 5.70 5.68 6.07 5.66All treatments b b a b 0.62 0.055Only 3 treatments B B A 0.70 0.034Only Hemicell 0.86 0.895Only Protexin B A 0.90 0.036Day 7 5.25 5.45 6.02 5.70All treatments B B A AB 0.68 0.012Only 3 treatments B B A 0.82 0.006Only Hemicell 1.03 0.752Only Protexin B A 1.07 0.015Day 14 5.62 5.80 5.80 5.53All treatments 0.55 0.210Only 3 treatments 0.63 0.422Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0114] Table 57 shows yolk yellowness (b* color) of eggs from laying hens fed beta- mannanase (Hemicell) and / or probiotics (Protexin), according to various storage times in a third trial.Table 57.Day 28 5.21 5.82 5.68 5.54All treatments B A A AB 0.64 0.003Only 3 treatments B A A 0.80 0.003Only Hemicell B A 1.16 0.006Only Protexin B A 1.08 0.026Day 35 5.55 5.80 5.74 5.50All treatments 0.51 0.102Only 3 treatments 0.54 0.156Only Hemicell b a 0.74 0.091Only Protexin 0.74 0.219Day 42 5.41 5.78 5.65 5.36All treatments B A AB B 0.51 0.007Only 3 treatments B A AB 0.53 0.011Only Hemicell B A 0.73 0.009Only Protexin B A 0.59 0.036Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.Example 3Oxidant and Antioxidant Assessments

[0115] For the instant example, assessment of oxidant and antioxidant levels birds was evaluated. The status in intestines and oviducts from laying hens in one of the four treatment groups described in Example 2 was evaluated.

[0116] To determine the levels of reactive oxygen species (ROS), the methodology described by Halliwell and Gutteridge (2007) was followed. The levels of non-homogeneous muscle lipid peroxidation were determined as levels of antibodies reactive to thiobarbituric acid (TBARS), measured by the absorbance of the red product at 532 nm, according to the method described by Ohkawa et al. (1978) and expressed as nmol MDA / mg of protein.

[0117] GST activity was measured using spectrophotometry at 340 nm, according to Habig et al. (1974). The mixture contained muscle homogenate supernatant as a test, 0.1 M potassium phosphate buffer (pH 7.4), 100 mM GSH and 100 mM CDNB, used as substrate. Enzyme activity was expressed as pmol / CDNB / mg protein. Protein thiols (PSH) were determined by the method described by Sedlak and Lindsay (1968), a technique that uses DTNB (5,5-dithiobis- 2- nitrobenzoic acid; Sigma). Protein thiols were measured by the sediment formed by the precipitated protein that material was resuspended, using ahomogenization buffer to determine the PSH content. The readings absorbances (405 nm) were performed using a spectrofluorometer (Biotek, Synergy HT).

[0118] Table 58 shows oxidant and antioxidant status in intestines and oviducts from laying hens fed beta-mannanase (Hemicell), and / or probiotics (Protexin). The variables evaluated included TBARS (thiobarbituric acid reactive substances; mmol MDA / mg proteins); GST (glutathione S-transferase; pmolCDNB / min / mg protein); and TSH (protein thios; pmol TSH / mg protein).Table 58.TreatmentsVariables Control P H H + P SE1P- value2IntestineTBARS 1.84A 1.79A 1.39AB 1.06B 0.12 0.018GST 975. IB 912.6B 914.9B 1251A 10.1 0.006TSH 1.49G 2.81A 1.78B 1.70B 0.04 <0.001OviductTBARS 0.70 0.907 0.77 0.81 0.08 0.440GST 861.2a 794.9a 741.7b 832.9a 15.3 0.061TSH 1.25 1.30 1.05 1.22 0.14 0.420Standard error.2Probability of treatment effect (model did not included the effect of storage time). Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.Example 4Performance Assessments

[0119] For the instant example, various performance were undertaken. The experimental units were randomly selected among the hens housed in the commercial farm (about 28,000 light-weight laying hens, 36 weeks old, Hyline W 36 lineage) and assigned to the treatments, that were i) control (basal diet, without supplementation); ii) diet supplemented with 300 g / ton of the P-mannanase Hemicell®; iii) diet supplemented with 50 g / ton of the probiotic additive Protexin®; iv) diet supplemented with 50 g / ton of probiotic additive and 300 g / ton of -mannanase (Hemicell+Protexin).

[0120] The basal feed was formulated according to the nutritional requirements for the genetic (Hy-Line, 2020). Corn and soybean meal were the main ingredients in the feed formulas. Feed and water were both provided ad libitum throughout the experimental period using nipple drinkers and gutter feeders.

[0121] The birds were housed in conventional sheds, arranged in an east-west direction, with concrete floors and masonry walls complemented with wire mesh to the ceiling. The shedwas equipped with side curtains, which were managed according to weather conditions to provide thermal comfort. Average maximum and minimum temperature and air relative humidity values recorded were 36 and 18.1 °C, and 94.7 and 35.8%, respectively. Only natural light illumination was used, allowing a lighting program of 16 hours and 30 minutes of light per day.

[0122] The instant example started when birds were 36 weeks old and lasted 144 days. Egg production was individually recorded at weeks 4, 8, and 12 in a subgroup of 120 cages with 4 birds each, corresponding to 30 replicates per treatment. All eggs produced in these subgroups were individually weighed and analyzed for the presence of feces in the shells, which was classified by the same observer through visual analysis as: absent (clean eggs), minor presence (e.g., presence of small spots; see Figure 1), or major presence (e.g., presence of large spots; see Figure 2).

[0123] Laying rate and egg mass were calculated considering all eggs (including non- marketable eggs) for each replicate (cage). The coefficient of variability was calculated for each cage considering the individual weight of all the eggs produced in each week. A similar procedure was adopted for evaluation of egg masses.

[0124] Blood samples were collected at the end of the project from 8 birds per treatment (random selection). Samples were processed and analyzed (Bio-Plus®, Biochemical Analyzer, Bioplus, Sao Paulo, Brazil) for total protein, albumin, uric acid, total cholesterol, triglycerides, glucose, alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase using commercial kits (Wiener Lab Group, Sao Paulo, Brazil) and semiautomatic analyzer.

[0125] Table 59 shows performance of laying hens fed diets supplemented with beta- mannanase (Hemicell) and / or probiotics (Protexin).Table 59.Treatments.. „ . , Hemicell Protexin .. „ P-Responses Control H + P SE12Laying rate (%)36 - 40 wk 85.31 C 93.71 B 83.25 C 97.66 A 0.11 <0.00141 - 44 wk 78.34 B 96.69 A 96.36 A 90.55 A 0.11 <0.00145 - 48 wk 91.05 B 92.59 AB 92.60 AB 95.90 A 0.09 <0.00136 - 48 wk 84.90 C 94.33 A 90.74 B 94.70 A 0.59 <0.001Weight of fresh eggs (g)36 - 40 wk 61.95 B 62.36 B 61.38 B 63.18 A 0.14 <0.00141 - 44 wk 61.28 C 63.09 A 61.98 B 62.79 A 0.15 <0.00145 - 48 wk 64.13 B 65.21 A 63.76 B 65.52 A 0.16 <0.00136 - 48 wk 62.47 B 63.55 A 62.37 B 63.83 A 0.09 <0.00162 wk (no treat.)362.53 B 64.40 A 64.10 A 64.28 A 0.13 0.013Coefficient of variability in egg weight (%)36 - 40 wk 5.944b 5.897 b 5.193 a 5.292 ab 0.014 0.07241 - 44 wk 7.152 B 5.625 A 5.750 A 5.734 A 0.018 0.00745 - 48 wk 7.088 B 5.397 A 5.608 A 5.405 A 0.019 0.00436 - 48 wk 6.728 B 5.640 Q 5.517 A 5.477 A 0.010 <0.00162 wk (no treat.)37.940 8.285 8.272 7.891 0.124 0.564'Slandard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%. treatments were not provided from wk 48 to 62. Thus, the last evaluation was performed after 14 wk without supplementation.

[0126] Table 60 shows egg component masses produced by hens fed diets supplemented with beta-mannanase (Hemicell) and / or probiotics (Protexin).Table 60.TreatmentsHemicell Protexin¥¥P-Responses Control H + P SE1 2Egg mass (g / hen / day)36 - 40 wk 5B2.85 B 58.44 A 51.10 B 61.70 A 0.67 <0.00141 - 44 wk 48.01 C 61.00 A 59.72 AB 56.86 B 0.39 <0.00145 - 48 wk 58.30 B 60.38 A 59.04 AB 62.83 A 0.63 <0.00136 - 48 wk 53.08 C 59.94 A 56.62 B 60.46 A 0.39 <0.001Yolk mass (g / hen / day)36 - 40 wk 15.89 B 17.36 A 14.57 B 18.46 A 0.19 <0.00141 - 44 wk 13.45 B 16.69 A 16.71 A 16.24 A 0.11 <0.00145 - 48 wk 14.81 C 15.58 B 15.67 B 16.70 A 0.18 <0.00136 - 48 wk 14.77 V 16.56 A 15.66 B 17.14 A 0.11 <0.001Albumen mass (g / hen / day36 - 40 wk 31.81 B 35.22 A 31.35 B 36.98 A 0.42 <0.00141 - 44 wk 29.54 B 38.05 A 36.95 A 34.72 A 0.23 <0.00145 - 48 wk 38.06 B 38.54 A 37.30 B 39.69 A 0.49 <0.00136 - 48 wk 33.09 C 37.26 A 35.18 B 37.12 A 0.24 <0.001Egg shell mass (g / hen / day)36 - 40 wk 5.14 B 5.78 A 5.21 B 6.26 A 0.07 <0.00141 - 44Wk 5.02 B 6.26 A 6.06 A 5.90 A 0.04 <0.00145 - 48 wk 5.42 C 6.26 A 6.06 B 6.44 A 0.06 <0.00136 - 48 wk 5.22 C 6.13 A 5.78 B 6.20 A 0.04 <0.0011Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0127] Table 61 shows occurrence of clean eggs or minor / major presence of feces (%) in eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin).Table 61.' Si andard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%. treatments were not provided from wk 48 to 62. Thus, the last evaluation was performed after 14 wk without supplementation.

[0128] Table 62 shows serum biochemistry and intestinal morphometry of laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin).Table 62.Villi area, pm2367039 288838 343272 450579 1798 0.670 Crypt depth, pm 224.9 186.4 227.6 202.1 4.96 0.397 Villi height : Crypt depth 6.839 B 7.245 AB 6.304 B 8.295 A 0.171 0.007Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.Example 5Egg Microstructure Assessments

[0129] For the instant example, microstructure of the eggs was evaluated. The eggshells obtained from eggs of laying hens in one of the four treatment groups described in Example 2 were evaluated.

[0130] Scanning electron microscopy analyzes were performed to assess the microstructure of the eggshells. For the evaluations, 20 shells from de last trial were used, in which fragments of approximately 5 cm2were removed and stored in falcon tubes. After this period, the samples were mounted on a stubs, coated with gold-palladium of 35 nm for 3 minutes (Sputter Coater - SCD 050 Balzers, Germany) and analyzed through a scanning electron microscope (JEOL 6060, Japan) at magnifications from 100 to 2000x. In the images, the thickness of mammillary layer, palisade layer, membrane layer, and vertical crystal layer were measured in three locations aleatorily chosen in each eggshell image. The size of column or mammillary bodies and the distance between mammillary knobs were also accessed, but five measurements were made in each image due to the larger variability among measurements.

[0131] Table 63 summarizes eggshell traits accessed through ultramicroscopy in eggs from laying hens fed beta-mannanase (Hemicell) and / or probiotics (Protexin) depending on storage times. Evaluations were obtained at the first phase (40 weeks), second phase (44 weeks), and third phase (48 weeks).Table 63.TreatmentsFirst Phase - 40 wkMammillary layer, pm 73.29 71.81 59.94 69.26 24.17 0.155Palisade layer, pm 238.6B 260.2AB 262.7A 254.0AB 22.50 0.035Mammill. -rPalisade, pm 317.8 332.0 315.4 315.7 28.10 0.319Membrane layer, pm 57.39 29.84 68.88 68.20 20.01 0.294Crystal layer, pm 28.39B 26.89B 26.80B 30.92A 19.32 0.429Mammillary bodies, pm 68.18B 65.82B 62.42B 71.34A 6.928 0.009Mam. Knobs distance, pm 83.89 82.34 68.13 78.30 21.41 0.148Second Phase - 44 wkMammillary layer, pm 50.86B 66.81A 65.56A 59.88A 12.03 0.003Palisade layer, pm 259.7B 284.3A 266.7 A 270.7 A 9.953 0.0131Standard error.2Probability of treatment effect. Means followed by different uppercase letters differ statistically at 5%, while lowercase letters were used to indicate differences at 10%.

[0132] Figure 3 shows ultramicroscopy images collected in a single eggshell at the first phase. Figure 4 shows ultramicroscopy images collected in a single eggshell at the second phase. Figure 5 shows ultramicroscopy images collected in a single eggshell at the third phase.

Claims

WHAT IS CLAIMED IS1. A method comprising a step of administering one or more supplements to a non-human animal, wherein the supplements comprise i) beta-mannanase and ii) a probiotic.

2. The method of claim 1, wherein the animal is an avian.

3. The method of claim 2, wherein the avian is selected from the group consisting of a chicken, a turkey, and a duck.

4. The method of claim 2, wherein the avian is a chicken.

5. The method of claim 3, wherein the chicken is a hen.

6. The method of claim 4, wherein the hen is a laying hen.

7. The method of claim 1, wherein the beta-mannanase is administered at a dose of 300 g / ton and wherein the probiotic is administered at a dose of 50 g / ton.

8. The method of claim 1, wherein the beta-mannanase and the probiotic provide a synergistic effect in improvement in health of the animal.

9. The method of claim 1, wherein the beta-mannanase is administered at a dose between 100 g / ton and 500 g / ton.

10. The method of claim 1, wherein the beta-mannanase is administered at a dose of 300 g / ton.

11. The method of any one of claims 1 to 5, wherein the supplement comprises a probiotic supplement comprising Enterococcus faecium.

12. The method of claim 11, wherein the Enterococcus faecium is National Collection of Industrial, Food and Marine Bacteria (NCIMB) culture number 11181.

13. The method of any one of claims 1 to 5, wherein the probiotic supplement comprises Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactobacillus plantarum, Lactobacillus rhamnosus, Bifidobacterium bifidum, Enterococcus faecium, and Streptococcus thermophiles.

14. The method of claim 12, wherein the probiotic supplement is administered at a dose of 50 g / ton.

15. The method of any one of claims 1 to 8, wherein the supplement is present in a feed for the animal.

16. The method of claim 8, wherein the feed is a free choice feed.

17. The method of any one of claims 1 to 8, wherein the administration provides an improvement in health of the animal.

18. The method of claim 17, wherein the improvement in health of the animal is an improvement in animal welfare.

19. The method of claim 17, wherein the improvement in health of the animal comprises a decrease in lesions on the animal.

20. The method of claim 17, wherein the improvement in health of the animal comprises an improvement in intestinal health in the animal.

21. The method of claim 17, wherein the improvement in health of the animal comprises prevention of intestinal dysbiosis in the animal.

22. The method of claim 17, wherein the improvement in health of the animal comprises a reduction in pro-inflammatory effect in the animal.

23. The method of claim 17, wherein the improvement in health of the animal comprises increased release of mannan-oiligosaccharides (MOS) in the animal.

24. The method of any one of claims 1 to 8, wherein the administration provides an improvement in egg quality of one of more eggs laid by the animal.

25. The method of claim 24, wherein the improvement in egg quality is indicated by a modification of a quality selected from the group consisting of weight, specific gravity, albumen height, albumen weight, yolk height, yolk length, yolk index, yolk weight, Haugh unit, shell weight, shell breaking strength, albumen pH, yolk pH, yolk color score, yolk lightness, yolk redness, yolk yellowness, chroma, and any combination thereof.

26. The method of any one of claims 1 to 8, wherein the administration provides an improvement in reactive oxygen species (ROS) in the animal.

27. The method of claim 26, wherein the improvement in ROS is indicated by a modification of thiobarbituric acid reactive substances (TBARS), glutathione S-transferase (GST), protein thios (TSH), and any combination thereof.

28. The method of any one of claims 1 to 8, wherein the administration provides an increase in egg laying rate of the animal.

29. The method of any one of claims 1 to 8, wherein the administration provides an increase in fresh egg weight for eggs laid by the animal.

30. The method of any one of claims 1 to 8, wherein the administration provides a reduction in coefficient of variability of egg weight for eggs laid by the animal.

31. The method of any one of claims 1 to 8, wherein the administration provides an increase in an egg component mass for eggs laid by the animal.

32. The method of any one of claims 1 to 8, wherein the administration provides an increase in occurrence of clean eggs laid by the animal.

33. The method of any one of claims 1 to 8, wherein the administration provides a modification in eggshell thickness in eggs laid by the animal.

Images