Experimentally segregated attenuated tick-borne encephalitis virus: altered behaviour after prolonged storage at subzero temperatures

In an attenuated variant of tick-borne encephalitis virus, western subtype (TBEV) with an ic+ sc s t e us character and lost virulence for monkeys, a distinct inhomogeneity together with a decrease in titre was observed after 8.7--9.6 years of storage at 4 and -14 (+/- 2) degrees C, with several interruptions due to failures in refrigeration systems. Proved by clonal analysis, the inhomogeneity concerned plaque polymorphism, increased thermo-resistance and degree of virulence for mice and monkeys. Marker studies of materials stored for up to 14.5 years yielded comparable findings. The possibility of the presence in the virus stocks in addition to a prevailing attenuated component also of minor component(s) having the advantage for survival under storage conditions, is discussed taking into account the intrinsic mutational capacity of the virus and known differences in resistance to environmental stresses between attenuated and virulent TBEV variants.