Isolation and Characterization of Clinical RSV Isolates in Belgium during the Winters of 2016–2018

Van der Gucht, Winke; Stobbelaar, Kim; Govaerts, Matthias; Mangodt, Thomas; Barbezange, Cyril; Leemans, Annelies; De Winter, Benedicte; Van Gucht, Steven; Caljon, Guy; Maes, Louis; De Dooy, Jozef; Jorens, Philippe; Smet, Annemieke; Cos, Paul; Verhulst, Stijn; Delputte, Peter L.

Viral neutralization by palivizumab was assessed with a conventional plaque reduction assay. Virus was incubated with a two-fold dilution series of palivizumab for 1h at 37 • C and then transferred to HEp-2 cells for 2h at 37 • C to allow infection of non-neutralized virus. Afterwards, the supernatant was replaced by DMEM-10 containing 0.6% Avicel ®® and incubated for three days until plaques were visible to the naked eye. Plaques were counted to determine the concentration of palivizumab that neutralizes the virus by 50%. Figure 7 shows that RSV-A clinical isolates BE/ANT-A7/17 and BE/ANT-A21/17 are 50% neutralized at lower palivizumab concentrations than most of the other clinical isolates and RSV A2, resulting in better neutralization than the other isolates. Remarkably, RSV A2 and RSV B1 neutralization was significantly different, with RSV-B strains having a higher sensitivity to palivizumab than RSV-A strains. Overall no significant differences were observed between RSV-B clinical isolates and the reference RSV B1 for palivizumab neutralization. Figure 7 shows that RSV-A clinical isolates BE/ANT-A7/17 and BE/ANT-A21/17 are 50% neutralized at lower palivizumab concentrations than most of the other clinical isolates and RSV A2, resulting in better neutralization than the other isolates. Remarkably, RSV A2 and RSV B1 neutralization was significantly different, with RSV-B strains having a higher sensitivity to palivizumab than RSV-A strains. Overall no significant differences were observed between RSV-B clinical isolates and the reference RSV B1 for palivizumab neutralization. with clinical isolates and reference strains that were pre-incubated for 1h with a palivizumab dilution series. Inoculum was replaced with DMEM-10 containing 0.6% Avicel ® and incubated for three days at 37°C. Afterwards, the cells were fixed, stained with palivizumab as primary antibody and goatanti-human conjugated with HRP, plaques were visualized with chloronapthol. Individual values are plotted as 2log ED50, data represents mean values ± SEM (n = 3). with clinical isolates and reference strains that were pre-incubated for 1h with a palivizumab dilution series. Inoculum was replaced with DMEM-10 containing 0.6% Avicel ®® and incubated for three days at 37 • C. Afterwards, the cells were fixed, stained with palivizumab as primary antibody and goat-anti-human conjugated with HRP, plaques were visualized with chloronapthol. Individual values are plotted as 2log ED50, data represents mean values ± SEM (n = 3).