Exogenous dietary lysozyme improves the growth performance and gut microbiota in broiler chickens targeting the antioxidant and non-specific immunity mRNA expression

<div><p>Supplementation of exogenous enzymes in chickens has been widely practiced, yet mechanisms responsible are not fully delineated. To investigate the effects of the dietary lysozyme on the growth performance and immunity of broiler chickens, a total of 120 one-day-old Ross 308 chicks were randomly allocated into four groups, each having three replicates (30 birds/group). The chicks were fed the starter (1–21 d) and grower (22–35 d) diets supplemented with 0 (control), 70 (LYZ70), 90 (LYZ90) and 120 (LYZ120) g of lysozyme 10%^® per ton of basal diet for five weeks. The results revealed significant improvement in the growth performance and gut environment. There were significant decreases (<i>P</i> < 0.05 or 0.01) in the harmful fecal <i>Coliform</i> and <i>Clostridia</i> and an increase (<i>P</i> ˂ 0.05) in the beneficial <i>Lactobacillus</i> in the lysozyme-supplemented groups, especially in LYZ90. Moreover, the mRNA expressions of Cu, Zn-superoxide dismutase (<i>SOD1</i>), glutathione peroxidase (<i>GSH-Px</i>), interferon-gamma (<i>IFN-γ</i>), interleukin-10 (<i>IL-10</i>), and interleukin-18 (<i>IL-18</i>) were upregulated in response to lysozyme supplementation. In comparison to control, LYZ90 fed birds had a significant increase (<i>P <</i> 0.01) in the <i>GSH-Px</i> gene expression that enhances the antioxidant status of the gut. Expression of the biomarkers involved in the gut non-specific immunity indicated significant increases in the mRNA expression of <i>INF-γ</i> (<i>P <</i> 0.001), <i>IL-10</i> (<i>P <</i> 0.001), and <i>IL-18</i> (<i>P <</i> 0.05) in LYZ90 group. Also, serum globulin levels were significantly elevated (<i>P</i> ˂ 0.05) in lysozyme-supplemented groups. Histologically, the intestinal villi length and crypts depth were also enhanced (<i>P</i> ˂ 0.05) by dietary lysozyme supplementation. In conclusion, supplementation of broiler chickens with exogenous lysozyme, especially at 90 g of lysozyme per ton of basal diet dose rate, improved the growth performance, gut antioxidant status, and nonspecific immunity of broiler chickens.</p></div

Exogenous dietary enzyme formulations improve growth performance of broiler chickens fed a low-energy diet targeting the intestinal nutrient transporter genes

<div><p>Diminishing the cost of broiler chicken diet is a critical issue in the poultry industry. Numerous studies were performed to achieve this pivotal objective by diet supplementation with alternative feed additives. In the current study, low-energy broiler rations were supplemented with different commercial multienzyme formulations to minimize the cost, and increase the digestibility and absorption of the digested macronutrients. Cobb Avian 48 broiler chicks (mixed sex, 1-d-old, <i>n</i> = 3120) were randomly allocated into six groups, and each group was subdivided into four replicates (130 birds per replicate). The birds were randomly allocated into a control group fed basal diet (CB); control group fed low-energy diet (CL); and birds fed low-energy diets supplemented with different enzyme formulations. The enzyme formulations used were Xylam 500^® (CLX group), Hemicell^® (CLH group), Avizyme^® (CLA group), and Megazyme^® (CLM group,) following the doses recommended by the manufacturers. The growth performance of CLA and CLH group birds was significantly improved when compared with CL. In comparison with CB, Avizyme^® significantly (<i>p</i> < 0.001) increased the intestinal <i>PEPT1</i>, <i>GLUT2</i>, <i>ACC</i>, and <i>IL-2</i> expression; PEPT1 facilitates the absorption of micronutrients. In conclusion, exogenous multienzyme complexes may be included in the low-energy diet to enhance the performance of broiler chickens (Avizyme^® ˃ Hemicell^® ˃ Megazyme^®), and reduce the diet cost by up-regulating the expression of intestinal nutrient transporter genes, and improving the immunity and serum biochemical parameters of broiler chickens.</p></div

Effect of lysozyme dietary supplementation on serum total protein (g/dl), albumin (g/dl), globulin (g/dl), ALT (U/L), AST (U/L), TAG (mg/dl), total cholesterol (mg/dl), creatinine (mg/dl), and uric acid (mg/dl) in broiler chickens.

<p>Effect of lysozyme dietary supplementation on serum total protein (g/dl), albumin (g/dl), globulin (g/dl), ALT (U/L), AST (U/L), TAG (mg/dl), total cholesterol (mg/dl), creatinine (mg/dl), and uric acid (mg/dl) in broiler chickens.</p

Components of Rogosa agar.

<p>Components of Rogosa agar.</p

RT-PCR validation of the <i>GLUT2</i> gene.

<p>Gene expression was analyzed in intestinal samples (<i>n</i> = 16). **<i>p</i> < 0.01 and ***<i>p</i> < 0.001 vs. CB. ⁺⁺⁺<i>p</i> < 0.001 vs. CL. ^xxx<i>p</i> < 0.001 vs. CLX. ^##<i>p</i> < 0.01 and ^###<i>p</i> < 0.001 vs. CLH. ^Θ<i>p</i> < 0.05, ^ΘΘ<i>p</i> < 0.01, and ^ΘΘΘ<i>p</i> < 0.001 vs. CLA. Statistical analysis was performed using one-way ANOVA and Tukey’s <i>post hoc</i> test for multiple comparisons. CB, control fed basal diet; CL, control fed low-energy diet; CLX, control fed low-energy diet containing Xylam 500^®; CLH, control fed low-energy diet containing Hemicell^®; CLA, control fed low-energy diet containing Avizyme^®; CLM, control fed low-energy diet containing Megazyme^®.</p

Primer sequences, target genes, and cycling conditions for SYBR green RT-PCR.

<p>Primer sequences, target genes, and cycling conditions for SYBR green RT-PCR.</p

Reverse transcription polymerase chain reaction (RT-PCR) validation of selected genes differentially expressed due to lysozyme (LYZ).

<p>Selected genes were <i>SOD1</i> = Cu, Zn-superoxide dismutase and <i>GSH-Px</i> = glutathione peroxidase. **<i>P</i> < 0.01 vs. control, ⁺<i>P</i> < 0.05 and ⁺⁺⁺<i>P</i> < 0.001 vs. LYZ90, and ^#<i>P</i> < 0.05 vs. LYZ120. Statistical analysis was done by One-way ANOVA, Tukey’s <i>post hoc</i> test multiple comparisons. Error bars represent SE.</p

Effect of lysozyme dietary supplementation on growth performance of broiler chickens.

<p>Effect of lysozyme dietary supplementation on growth performance of broiler chickens.</p

Composition of the experimental starter and finisher diets.

<p>Composition of the experimental starter and finisher diets.</p

Histological examination of jejunum samples stained with H&E.

<p>(A) control (×40), (B) LYZ70 (×40), (C) LYZ90 (×100), and (D) LYZ120 (×40). Large arrows point to intestinal villi. Small arrows point to intestinal crypts.</p