Dataset Information

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-?, IFN-?, 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.

SUBMITTER: Sureshkumar S

PROVIDER: S-EPMC6591341 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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

Sureshkumar Shanmugam S Jung Sun Keun SK Kim Dongjun D Oh Keon Bong KB Yang Hyeon H Lee Hwi Cheul HC Jin Jo Yong JY Sun Lee Hae LH Lee Sukchan S Byun Sung June SJ

3 Biotech 20190624 7

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. Admini ...[more]

PMID: 31245246

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Project description:BackgroundIntimin is an important virulence factor involved in the pathogenesis of enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic Escherichia coli (EHEC). Both pathogens are still important causes of diarrhea in children and adults in many developing and industrialized countries. Considering the fact that antibodies are important tools in the detection of various pathogens, an anti-intimin IgG2b monoclonal antibody was previously raised in immunized mice with the conserved sequence of the intimin molecule (int388-667). In immunoblotting assays, this monoclonal antibody showed excellent specificity. Despite good performance, the monoclonal antibody failed to detect some EPEC and EHEC isolates harboring variant amino acids within the 338-667 regions of intimin molecules. Consequently, motivated by its use for diagnosis purposes, in this study we aimed to the cloning and expression of the single-chain variable fragment from this monoclonal antibody (scFv).FindingsAnti-intimin hybridoma mRNA was extracted and reversely transcripted to cDNA, and the light and heavy chains of the variable fragment of the antibody were amplified using commercial primers. The amplified chains were cloned into pGEM-T Easy vector. Specific primers were designed and used in an amplification and chain linkage strategy, obtaining the scFv, which in turn was cloned into pAE vector. E. coli BL21(DE3)pLys strain was transformed with pAE scFv-intimin plasmid and subjected to induction of protein expression. Anti-intimin scFv, expressed as inclusion bodies (insoluble fraction), was denatured, purified and submitted to refolding. The protein yield was 1 mg protein per 100 mL of bacterial culture. To test the functionality of the scFv, ELISA and immunofluorescence assays were performed, showing that 275 ng of scFv reacted with 2 mg of purified intimin, resulting in an absorbance of 0.75 at 492 nm. The immunofluorescence assay showed a strong reactivity with EPEC E2348/69.ConclusionThis study demonstrated that the recombinant anti-intimin antibody obtained is able to recognize the conserved region of intimin (Int388-667) in purified form and the EPEC isolate.

Dataset Information

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

Publications

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

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