Abstract
Domain III of the envelope protein (EDIII) is the major target of flavivirus neutralizing antibody. To date, little is known regarding antibody-mediated neutralization of Tembusu virus (TMUV), a novel flavivirus emerging in duck in 2010. Here, a novel monoclonal antibody (MAb), designated 12F11, was prepared by immunization of mice with recombinant EDIII (rEDIII) protein. Using virus neutralization test, 12F11 in undiluted ascites neutralized the TMUV infectivity to induce the development of cytopathic effects in BHK-21 cells, indicating that 12F11 exhibits a neutralizing activity. The neutralizing activity of 12F11 was confirmed by plaque reduction neutralization test, in which 12F11 reduced significantly the number of plaques produced by TMUV in BHK-21 cells. Western blot analyses of a series of truncated rEDIII proteins showed that the epitope recognized by 12F11 includes amino acids between residues 8 and 77 of EDIII protein. Function analysis demonstrated that 12F11 neutralizes TMUV infection at virus adsorption and at a step after adsorption to a certain extent. The present study provides an important step towards elucidating antibody-mediated neutralization of TMUV.
Highlights
Tembusu virus (TMUV), one of the members in the genus Flavivirus, was first isolated from mosquitoes in 1955 in Malaysia [1]
The EDIII protein of TMUV Y was predicted to comprise 109 amino acids, which corresponded to residues 298–406 in the E protein (Figure 1A)
Alignment of the EDIII protein of TMUV Y with those of DENV, JEV, and WNV whose three-dimensional structures are known revealed the detectable sequence conservation, which allowed a prediction of the amino acids making up seven β-strands (Figure 1B)
Summary
Tembusu virus (TMUV), one of the members in the genus Flavivirus, was first isolated from mosquitoes in 1955 in Malaysia [1]. The virus caused explosive outbreaks of a severe disease in ducks in eastern China at the end of spring and the beginning of summer of 2010. In the six months, the disease spread rapidly to other regions with intensive duck productions, including southern. Multiple strategies have been employed for generating effective vaccines [5,6,7,8,9,10,11]. A recent study showed that TMUV mediates antibody-dependent disease severity in mice [12]. It is crucial to define the epitopes which induce the potent neutralizing antibodies for development of a safe and effective TMUV vaccine
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