Oral Immunization with Recombinant Lactobacillus acidophilus Expressing espA-Tir-M Confers Protection against Enterohemorrhagic Escherichia coli O157:H7 Challenge in Mice.

Ruqin Lin,Hongying Fan,Bo Zhu,Xianbo Wu,Siqin Duan,Beiguo Long,Yiduo Zhang,Yuhua Wu,Yawen Li

doi:10.3389/fmicb.2017.00417

Abstract

Enterohemorrhagic Escherichia coli O157:H7 (EHEC O157:H7) causes hemorrhagic colitis and the formation of characteristic attaching and effacing (A/E) lesions in humans. Given the severe sequelae of EHEC O157:H7 infection, it is critical to develop effective vaccines for human use. However, for achieving this goal many hurdles need to be addressed, such as the type or subset of antigens, adjuvant, and the delivery route. We developed a candidate vaccine by inserting the bivalent antigen espA-Tir-M composed of espA and the Tir central domain into Lactobacillus acidophilus. The recombinant L. acidophilus (LA-ET) was safe in a cell model and excluded EHEC O157:H7 from LoVo cells at rates of nearly 94 and 60% in exclusion and competition assays, respectively. LA-ET inhibited the induction of A/E lesions by EHEC O157:H7 cells in vitro. Oral immunization with LA-ET induced higher levels of specific mucosal and systemic antibody responses in mice. Moreover, LA-ET enhanced interferon-γ and interleukin-4 and -10 production, which was associated with mixed helper T (Th1/Th2) cell responses, and protected against EHEC O157:H7 colonization and infection in mice at a rate of 80%. Histopathological analyses revealed that orally administered LA-ET reduced or inhibited A/E lesions and toxin-induced systemic injury. These findings demonstrate that LA-ET induces both humoral and cellular immune responses in mice and is therefore a promising vaccine against EHEC O157:H7 infection.

Highlights

Enterohemorrhagic Escherichia coli O157:H7 (EHEC O157:H7) is a zoonotic enteric pathogen that causes hemorrhagic colitis and hemolytic uremic syndrome (HUS) in humans (Remis et al, 1984; Armstrong et al, 1996)
The espA-Tir central domain (Tir-M) fusion gene was amplified by overlap extension PCR, yielding the expected 816-bp product, which was visualized by agarose gel electrophoresis
The fragment was cloned into the pMG36e vector, and the recombinant pMG36e-espA-Tir-M plasmid was transformed into L. acidophilus cells

Summary

Introduction

Enterohemorrhagic Escherichia coli O157:H7 (EHEC O157:H7) is a zoonotic enteric pathogen that causes hemorrhagic colitis and hemolytic uremic syndrome (HUS) in humans (Remis et al, 1984; Armstrong et al, 1996). Two vaccines are used for the same purpose in cattle, but neither has been approved for humans (Larrie-Bagha et al, 2013). For achieving this goal many hurdles need to be addressed, such as the type or subset of antigens, adjuvant, and the delivery route. The T3SS proteins, espA and Tir play key roles in EHEC adhesion, pedestal formation, and pathogenicity. They may serve as immunogens for the disruption of bacterial-host cell interaction and prevent the formation of A/E lesions

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Discussion

Conclusion