microbiology, Vol 141, 1993-2002, Copyright © 1995 by Society for General Microbiology
Attenuated typhoid vaccine Salmonella typhi Ty21a: fingerprinting and quality control
AJ McKenna, JA Bygraves, MC Maiden and IM Feavers
Division of Bacteriology, National Institute for Biological Standards and Control, Potters Bar, Hertfordshire, UK.
Live attenuated vaccines, developed with molecular genetical techniques,
require new approaches for their quality control. To develop novel quality
control tests that enhanced and extended existing procedures, the
attenuated vaccine strain Salmonella typhi Ty21a and its parent strain Ty2
were characterized by pulsed-field gel electrophoresis (PFGE) and direct
nucleotide sequence analysis. Mutant and parent strains were distinguished
using fingerprints generated by the resolution on PFGE of chromosomal DNA
digested with each of the enzymes SfiI, SpeI or XbaI. These fingerprints
were stable through multiple in vitro passages of the vaccine strain and
were identical from one batch of vaccine to another. It was also possible
to distinguish between the mutant and parent strains by direct nucleotide
sequence analysis of the galE gene. This analysis identified two base
changes in the gene from strain Ty21a: a single base deletion causing a
frameshift that would result in a truncated gene product, accounting for
the galE phenotype; and a transition that eliminated an AluI restriction
site. The consequent change in the AluI fingerprint of the galE gene in
strain Ty21a provided a rapid, PCR-based alternative to the use of
differential media or biochemical assays for the identification of the
vaccine strain.
