Phenotypic and functional characterization of T-cells and in vitro replication of FMDV serotypes in bovine lymphocytes

Abstract

Peripheral blood mononuclear cells (PBMCs) from unvaccinated and vaccinated bovine calves were employed to study the alteration of different T-cell subpopulations, functional competence and detection of foot-and-mouth disease virus (FMDV) at different hours post in vitro infection (hpi) with FMDV serotype either O or A or Asia1. The alteration in T-lymphocyte subpopulations was analyzed by flow cytometry analysis using T-cell subpopulation specific monoclonal antibodies at various hpi. All the three FMDV serotypes down-regulate boCD4+ and boCD8+ T-cells up to 48hpi whereas down-regulation of boWC1+ T-cells was observed up to 48hpi with FMDV serotype O only from unvaccinated calves. In contrast, lymphocytes from vaccinated animals demonstrated significant up-regulation of boCD4+, boCD8+ and boWC1+ T-cells following exposure to FMDV serotype either O or A or Asia1. The lymphoproliferative test for functional competence of PBMCs using PHA or inactivated FMDV serotype either O or A or Asia1 antigens, revealed that PHA driven lymphoproliferative response (LPR) was significantly suppressed at 24, 48 and 72hpi in unvaccinated animals. Homotypic as well as heterotypic FMDV antigen specific LPRs were suppressed in PBMCs of unvaccinated animals infected with FMDV serotype either O or A or Asia1 and stimulated with FMDV antigens. FMDV was demonstrated by reverse transcription-polymerase chain reaction (RT-PCR) and 3A-non-structural protein (NSP) expression in PBMCs infected in vitro with FMDV serotype either O or A or Asia1 at different hpi. Using RT-PCR, the amplified product revealed 1D gene specific product of FMDV serotype O (1301bp) or A (866bp) or Asia1 (914bp) in lymphocytes of both unvaccinated and vaccinated calves at different hpi with FMDV serotype either O or A or Asia1, respectively. Likewise, the 3A-NSP mAb based flow cytometry revealed significant increase in 3A-NSP expression in lymphocytes of unvaccinated and vaccinated calves at different hpi with FMDV serotype either O or A or Asia1. In conclusion, the present study provides significant evidence that FMDV infects PBMCs in vitro and leads to transient immuno-suppression of boCD4+ and boCD8+ T-cells.