Iron-uptake systems of Vibrio anguillarum

Mackie, Carol (1987) Iron-uptake systems of Vibrio anguillarum. PhD thesis, University of Glasgow.

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This thesis is centrally concerned with the characterisation of iron-uptake systems in Vibrio strains isolated from salmonid fish and oysters and the possible role of these systems in pathogenesis. Altogether 23 strains were examined, 13 were Vibrio anguillarum, 3 were Vo tubiashi, 1 was Vo alginolyticus and 6 were unclassified Vibrio strains. Of the 23 strains, only three contained a plasmid of similar molecular weight (45-50 Mdal) to the virulence plasmid pJMI of Vo anguillarum strain 775. Three of the 23 strains could not grow in the presence of transferrin indicating that there was no correlation between plasmid carriage and growth under conditions of iron limitation. In order to facilitate the serological grouping and further characterise these strains, the LPS of ten V. anguillarum strains were compared by SDS-PAGE and immunoelectroblotting with rabbit antiserum raised against the cell envelope of strain 775. LPS from three strains, strains 1197, 4979 and 775, reacted strongly with this antiserum. All three strains were of serotype J-0-3. Absorbed antiserum against strain 775 which did not react with LPS on immunoelectroblotting still reacted with a further component in the cell envelope of strains 775, 1197 and 4979. This envelope component represents an additional antigen, to the major antigen of LPS, which is specific to serotype J-0-3. The iron-uptake systems of these ten V. anguillarum strains were then investigated by siderophore analysis, 55Fe-uptake and detection plus immunoanalysis of cell envelope proteins associated with iron limitation. Cell envelope proteins produced in the presence of transferrin were detected by an EDTA extraction procedure and PAGE analysis, V. anguillarum strains NCMB6, 636, 827, 4979, 91079 and 2981 produced one additional protein (Mr = 68-73,000), strains 775, 1197 and 5679 produced two (Mr = 73,000 and 78-79,000 respectively) and strain 1445 produced three (Mr = 68, 72 and 73,000) " Antiserum produced in rabbits to cell envelope fractions of V. anguillarum 775 grown under conditions of iron limitation was absorbed with envelope fractions from cells grown in iron-replete conditions" With this absorbed antiserum, which contained antibodies to the two cell envelope proteins associated with iron limitation, the ten strains were examined on nitrocellulose paper after immunoelectroblotting. Antigenic similarity was seen between proteins associated with iron limitation in all strains. One protein cross-reacted with the absorbed antiserum in strains NCMB6, 636, 827, 1445, 91079, 2981 and 4979 and two proteins cross-reacted in strains 1197 and 5679 indicating that two possible iron-uptake systems existed in V. anguillarum. The ten strains of V. anguillarum produced three different types of iron-binding compounds when cultured under different conditions. These were (1) a common phenolate-type produced by all ten strains, (2) an hydroxamate-type produced by three strains when grown in Tris- succinate medium (TSM) and (3) a second phenolate-type produced by strains 775, 1197 and 4979, all of which contained a plasmid of 45-50 Mdal molecular weight. The relative affinities of these siderophores, determined by competition for 55Fe was; Second phenolate-type> Hydroxamate-type > Common phenolate-type. However, under these conditions, none removed iron from desferal or purified aerobactin. Experimental infections of rainbow trout (Salmo gairdneri) were carried out with V. anguillarum strains NCMB6, 775 and 91079, strains which produced different combinations of siderophores in vitro. Only the common-type siderophore was detected in the kidney and spleen of fish infected with strains NCMB6 and 91079. The hydroxamate-type siderophore produced vitro by strain NGMB6 was not detected in vivo. However, in the kidney of fish infected with strain 775, both the common and second phenolate-type siderophores were detected, suggesting that the second uptake system was required by strain 775 in vivo and that the iron-uptake system based on the common phenolate-type siderophore was apparently defective. It was concluded that two independent iron-uptake systems existed in V. anguillarum, one plasmid-mediated, the other chromosomal- mediated and that these systems are produced during infections of rainbow trout.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Dr. T.H. Birkbeck
Keywords: Microbiology.
Colleges/Schools: College of Medical Veterinary and Life Sciences
Supervisor's Name: Supervisor, not known
Date of Award: 1987
Depositing User: Enlighten Team
Unique ID: glathesis:1987-73094
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 14 Jun 2019 08:56
Last Modified: 14 Jun 2021 14:57

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