Applications of Next-Generation Sequencing Technologies to Diagnostic Virology

Barzon, Luisa; Lavezzo, Enrico; Militello, Valentina; Toppo, Stefano; Palù, Giorgio

Intrinsic genetic instability of RNA viruses may lead to the accumulation of virulent revertants during manufacture of live viral vaccines, requiring rigorous quality control to ensure vaccine safety. High throughput deep sequencing methods have been proposed as tools for monitoring genetic consistency of live viral vaccines. Deep sequencing was used to analyze lots of oral poliovirus vaccine and the detected neurovirulence mutations were identical to the mutation detected with the standard method based on PCR and restriction enzyme cleavage [132] . Patterns of mutations present at a low level in vaccine preparations were characteristic of seed viruses used for their manufacture and could be used for identification of individual batches [132] . Deep sequencing was also used to examine eight live-attenuated viral vaccines, i.e., trivalent oral poliovirus, rubella, measles, yellow fever, varicella-zoster, multivalent measles/mumps/rubella, and two rotavirus live vaccines [133] . The method allowed identification of, not only mutations and minority variants relative to vaccine strains, but also sequences of adventitious viruses from the producer avian and primate cells. The results were in agreement with those obtained by using a panmicrobial microarray [133] .