Detection of Neutralizing Antibodies to Tembusu Virus: Implications for Infection and Immunity

Junfeng Lv; Lixin Yang; Shenghua Qu; Runze Meng; Qingxiangzi Li; Huicong Liu; Xiaoyan Wang; Dabing Zhang

doi:10.3389/fvets.2019.00442

Detection of Neutralizing Antibodies to Tembusu Virus: Implications for Infection and Immunity

Front Vet Sci. 2019 Dec 10:6:442. doi: 10.3389/fvets.2019.00442. eCollection 2019.

Authors

Junfeng Lv¹, Lixin Yang¹, Shenghua Qu¹, Runze Meng¹, Qingxiangzi Li¹, Huicong Liu¹, Xiaoyan Wang¹, Dabing Zhang¹

Affiliation

¹ Key Laboratory of Animal Epidemiology and Zoonosis of Ministry of Agriculture, College of Veterinary Medicine, China Agricultural University, Beijing, China.

Abstract

Neutralizing antibodies are the key mediators of protective immune response to flaviviruses after both infection and vaccination. Plaque reduction neutralization test (PRNT) is considered the "gold standard" for measurement of the immunity. To date, little is known regarding neutralizing antibody response to Tembusu virus (TMUV), a novel flavivirus emerging in ducks in 2010. Here, we developed a PRNT for detection of TMUV neutralizing antibodies. Following optimization and validation, the PRNT was applied to test serum samples from different flocks of ducks. Using sera prepared in experimental conditions, the levels of 50% end point titer (neutralizing dose, ND₅₀) generated from positive sera (5,012-79,433) were significantly higher than those from mock-infected sera (10 to 126), indicating that the test can be used in the detection of TMUV-specific neutralizing antibodies. Dose-dependent efficacy test of a cell-derived 180th passage of a plaque-purified virus of the PS TMUV isolate (PS180) in combined with immunization-challenge experiments revealed that ND₅₀ titer of ~1,258 is the minimum capable of providing adequate protection against challenge with virulent TMUV. In the investigation of serum samples collected from three flocks infected by TMUV and four flocks vaccinated with a licensed attenuated vaccine (the 120th passage virus), ND₅₀ titers peaked at 1 week after both disease onset (7,943-125,893) and vaccination (3,612-79,432), and high levels of ND₅₀ titer were detected in sera collected at 15 weeks after disease onset (5,012-63,095) and 17 weeks after vaccination (3,981-25,119). Together these findings demonstrated that spontaneous and experimental infections by TMUV and vaccination with the licensed TMUV attenuated vaccine elicit high, long-lasting neutralizing antibodies. The highest ND₅₀ titer of neutralizing antibodies elicited by PS180 was determined to be 3,162, suggesting that attenuation of TMUV by more passages has a dramatic impact on the neutralizing antibody response of the virus.

Keywords: TMUV attenuated vaccine; TMUV infection; Tembusu virus; duck; humoral immune response; neutralizing antibody; plaque reduction neutralization test.