Expression of recombinant human lysozyme in transgenic chicken promotes the growth of Bifidobacterium in the intestine and improves postnatal growth of chicken.

Hai Wang,Kejun Wang,Ling Lian,Kun Yu,Zhichen Cao,Zhengxing Lian,Hongping Wu

doi:10.1186/s13568-016-0280-2

Abstract

Lysozyme is one kind of antimicrobial proteins and often used as feed additive which can defend against pathogenic bacteria and enhance immune function of animals. In this study, we have injected the lentiviral vector expressing recombinant human lysozyme (rhLZ) gene into the blastoderm of chicken embryo to investigate the effect of recombinant human lysozyme on postnatal intestinal microbiota distribution and growth performance of chicken. Successfully, we generated 194 transgenic chickens identified by Southern blot with a positive transgenic rate of 24%. The average concentration of rhLZ was 29.90 ± 6.50 μg/mL in the egg white. Lysozyme in egg white of transgenic chickens had a significantly higher antibacterial activity than those of non-transgenic chickens by lysoplate assay (P < 0.05). The feces of transgenic and non-transgenic chickens were collected and five types of bacteria (Lactobacillus, Salmonella, Bifidobacterium, Staphylococcus aureus and Escherichia coli) were isolated and cultured to detect the impact of rhLZ on gut microbiota. Among the five bacteria, the number of Bifidobacterium in the intestine of those transgenic was significantly increased (P < 0.05). Moreover, the growth traits of the transgenic and non-transgenic chickens were analyzed. It was found that the 6-week shank length, 6-week weight and 18-week weight of transgenic chickens were significantly increased than that of non-transgenic chickens. The results demonstrated that rhLZ-transgenic chicken could promote the growth of Bifidobacterium in the intestine and improve the postnatal growth of chicken.

Highlights

Lysozyme (EC.3.2.1.17) widely exists in eukaryotes and prokaryotes to protect organism from microbial invasion
Transfection efficiency of eukaryotic expression vectors and expression of enhanced green fluorescent protein (EGFP) in transfected 293FT cells Eukaryotic expression vectors pChLZ-IRES2-EGFP and pHmLZ-IRES2-EGFP were constructed (Fig. 1). 293FT cells transfected with pChLZ-IRES2-EGFP showed highest expression of green fluorescent protein at 48 h post transfection (Fig. 2a, b), 293FT cells transfected with pHmLZIRES2-EGFP showed medium expression (Fig. 2c, d), and 293FT cells transfected with pIRES2-EGFP showed low expression
Consistent to these findings, we found that egg white of transgenic chickens exhibited a stronger antibacterial activity by agar diffusion test in this study

Summary

Introduction

Lysozyme (EC.3.2.1.17) widely exists in eukaryotes and prokaryotes to protect organism from microbial invasion. The c-type lysozyme exists ubiquitously in many organisms including viruses, bacteria, plants, insects, fishes, reptiles, birds, and mammals (Maga et al 2006a; Jolles 1996). Human lysozyme (hLZ), one of c-type lysozymes, is a positively charged molecule that consists of a polypeptide of 130 amino acid residues (Booth et al 1997; Yu et al 2013). It is widely distributed in body fluids, such as tears, saliva, and blood and plays an important role in host defenses (Jolles and Jolles 1984).

Methods

Results

Conclusion