Characterization of PM20C2, A TadD- Pasteurella muliocida Mutant
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Authors
Cotey, Alicia D.
Issue Date
2003
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
A mutant strain of Pasteurella multocida, PM20C2, which contains a transposon
disruption in the TadD gene, was characterized using genetic complementation studies
and an in vitro/in vivo competition assay to assess its potential for use in a live vaccine.
Complementation was achieved by incorporating either the wildtype P. multocida TadCD
genes or the wildtype Actinobacillus actinomycetemcomitans TadCD genes into the
pNF2176 plasmid. These complemented mutants were then used to infect mice and the
resultant LD50 values were compared to the LD50 values of mice infected with the uncomplemented PM20C2 mutant, the wildtype TF5 P. multocida strain,or the PM20C2
mutant complemented with an empty pNF2176 vector plasmid. Neither TadCD from P.
multocida nor that from A. actinomycetemcomitans successfully restored virulence. It is
likely that the transposon disrupted the expression of genes downstream from its
insertion in TadD. The in Vitro/in vivo competition assay involved creating a mixture of
equivalent concentrations (cfu/ml) of PM20C2 and TF5. This was then used to either
infect a mouse model or a falcon tube of broth. The output ratio was determined and
entered, along with the input ratio, into the Competitive Index equation. The Competitive
Index equation gives a numerical value to the fitness of the mutant when placed in
competition with the wildtype. When compared to the wildtype, the PM20C2 mutant
was near equally fit in vivo and less fit in vitro. Possible complementation by the
presence of the wildtype, TF5, may explain the discrepancy in viability of the PM20C2
mutant in the competition assays vs. Signature Tagged Mutagenesis. From the results
obtained, it is still inconclusive as to whether PM20C2 would be a good candidate for a
live vaccine·.
Description
v, 28 p.
Citation
Publisher
Kalamazoo College
License
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