Plaque purification
Plaque assay was used to obtain purified virus clones for selected
samples showing unequivocal results with pathotype-specific RT-qPCRs.
Briefly, different virus dilutions were incubated on confluent LMH cell
monolayers cultured in six-well plates overnight at 37°C. Then, virus
inoculum was removed, cell layers washed carefully twice with sterile
PBS and overlaid with modified Eagle’s medium containing 1.8 % agar.
Infected cells were incubated at 37 °C and 55% humidity for three days.
Finally, selected plaques were picked and propagated on LMH cells as
described above for further characterization.