Affiliation:
1. Departamento de Patologı́a (Animal Sanidad Animal), Unidad de Microbiologı́a e Inmunologı́a,1 and
2. Departamento de Biologı́a Celular y Anatomı́a,2 Universidad de León, 24007 León, Spain
Abstract
ABSTRACT
The
aroA
gene of
Aeromonas hydrophila
SO2/2, encoding 5-enolpyruvylshikimate 3-phosphate synthase, was cloned by complementation of the
aroA
mutation in
Escherichia coli
K-12 strain AB2829, and the nucleotide sequence was determined. The nucleotide sequence of the
A. hydrophila aroA
gene encoded a protein of 440 amino acids which showed a high degree of homology to other bacterial AroA proteins. To obtain an effective attenuated live vaccine against
A. hydrophila
infections in fish, the
aroA
gene was inactivated by the insertion of a DNA fragment containing a kanamycin resistance determinant and reintroduced by allelic exchange into the chromosome of
A. hydrophila
AG2 by means of the suicide vector pSUP202. The
A. hydrophila
mutant AG2
aroA
::Ka
r
was highly attenuated when inoculated intraperitoneally into a rainbow trout, with a 50% lethal dose of >2 × 10
8
CFU. The mutants were not recoverable from the internal organs after 48 h postinoculation. Immunohistochemical studies demonstrated that immunopositive materials, but not whole cells, reacting with a polyclonal antiserum against
A. hydrophila
were present in the kidney and spleen 9 days postinjection. Vaccination of rainbow trout with the AroA mutant as a live vaccine conferred significant protection against the wild-type strain of
A. hydrophila.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
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