Affiliation:
1. Dipartimento di Biologia Cellulare e dello Sviluppo, Fondazione Istituto Pasteur-Cenci Bolognetti, Università di Roma “La Sapienza,” 00185 Rome, Italy
Abstract
ABSTRACT
We have constructed and analyzed a group of
Shigella flexneri
5 auxotrophic mutants. The wild-type strain M90T was mutagenized in genes encoding enzymes involved in the synthesis of (i) aromatic amino acids, (ii) nucleotides, and (iii) diaminopimelic acid. In this way, strains with single (
aroB
,
aroC
,
aroD
,
purE
,
thyA
, and
dapB
) and double (
purE aroB
,
purE aroC
,
purE aroD
,
purE thyA
) mutations were obtained. Although the Aro mutants had the same nutritional requirements when grown in laboratory media, they showed different degrees of virulence in vitro and in vivo. The
aroB
mutant was not significantly attenuated, whereas both the
aroC
and
aroD
strains were severely attenuated.
p
-Aminobenzoic acid (PABA) appeared to be the main requirement for the Aro mutants’ growth in tissue culture. Concerning nucleotides, thymine reduced the pathogenicity, whereas adenine did not. However, when combined with another virulence-affecting mutation, adenine auxotrophy appeared to potentiate that mutation’s effects. Consequently, the association of either the
purE
and
aroC
or the
purE
and
aroD
mutations had a great effect on virulence as measured by the Sereny test, whereas the
purE aroB
double mutation appeared to have only a small effect. All mutants except the
dapB
strain seemed to move within a Caco-2 cell monolayer after 3 h of infection. Nevertheless, the auxotrophs showing a high intracellular generation time were negative in the plaque assay. Knowledge of each mutation’s role in attenuating
Shigella
strains will provide useful tools in designing vaccine candidates.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Cited by
67 articles.
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