Oral administration of Lactobacillus reuteri expressing a 3D8 single-chain variable fragment (scFv) enhances chicken growth and conserves immune homeostasis

Abstract

The present study was aimed to investigate the effects of 3D8 scFv-secreting Probiotic Lactobacillus reuteri (L. reuteri) on growth performance, inflammatory responses, and intestinal microbial flora in chickens. To this end, a total of 14 healthy wild-type chickens were divided into two experimental groups. Each group was orally administrated with a daily dose of 10⁹ colony-forming units (CFU) of 3D8 scFv-producing L. reuteri or wild-type (WT) for 35 days. Administration of L. reuteri/3D8 scFv significantly improved the body weight of chickens when compared to L. reuteri/WT group. The bacterial taxonomic composition of the fecal microbiota was determined by pyrosequencing of 16S rRNA gene amplicons. Firmicutes, Actinobacteria, and Proteobacteria were dominant phyla in two experimental groups. However, in 3D8 L. reuteri treatment groups at genus level, the Lactobacillus was highly abundant, being represented by 18.12%. In addition, serum levels of primary cytokines such as IL-6, IL-8, TNF-α, IFN-γ, IL-4, and IGF1 were markedly reduced in the probiotic L. reuteri 3D8 group. In summary, our results indicate that the administration of L. reuteri expressing 3D8 scFv has a modulatory effect on inflammatory responses, improves weight gain while not affecting the common microbial composition of the chicken intestine.

Keywords: 3D8 scFv; Chicken; Lactobacillus reuteri; Oral administration receive.

Fig. 1

Cloning strategy for cell wall secretion of 3D8 scFv via the pSLP-LDH expression vector in Lactobacillus reuteri (a). L. reuteri designed to secrete 3D8 scFv (b). P_LDH Lactobacillus dehydrogenase promoter, Ss SlpA secretion signal, Pro A protein A tag to prevent fusion of 3D8 scFv with the cell wall-anchoring domain of PrtP (anchor sequence). For 3D8 scFv protein expression, a codon-optimized 3D8 scFv gene (975 bp) was cloned into the pSLP-LDH vector (c)

Fig. 2

The expression level of 3D8 scFv protein in recombinant L. reuteri and recombinant L. paracasei ATCC 334 strains. The expression level of 3D8 scFv in transformed L. reuteri was determined with SYBR Premix Ex Taq by real-time RT-PCR

Fig. 3

Effect of L. reuteri secreting 3D8 scFv on growth performance traits in chickens. Statistical comparisons were made between chickens that received WT or 3D8 scFv-expressing L. reuteri administration once/day for 35 days. The results are expressed as the mean ± SE or SD

Fig. 4

Effects of L. reuteri secreting 3D8 scFv on serum cytokine levels in chickens. Statistical comparisons were made between chickens that received WT or 3D8 scFv-expressing L. reuteri once/day for 35 days. The results are expressed as mean ± SE or SD

Fig. 5

Bacterial taxonomic composition for the fecal microbiota of the chickens that received L. reuteri dietary treatment at the phylum level, in which each bar in the stacked bar charts represents the classifications of the total sequences

Fig. 6

Bacterial taxonomic composition from the fecal microbiota of the L. reuteri dietary supplementation groups at the genus level, in which each bar in the stacked bar charts represents the classifications of the total sequences