5 The role of quorum-sensing in the virulence of Pseudomonas ...
5 The role of quorum-sensing in the virulence of Pseudomonas ...
5 The role of quorum-sensing in the virulence of Pseudomonas ...
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<strong>Pseudomonas</strong> aerug<strong>in</strong>osa PA14 (UCBPP-PA14) is a wild-type reference stra<strong>in</strong> that<br />
has been used extensively to test <strong>the</strong> contribution <strong>of</strong> <strong>virulence</strong> factors to disease.<br />
<strong>The</strong> P. aerug<strong>in</strong>osa stra<strong>in</strong> PAO1, ano<strong>the</strong>r wild-type reference stra<strong>in</strong>, has been noted<br />
to be less virulent than P. aerug<strong>in</strong>osa PA14 <strong>in</strong> a number <strong>of</strong> models. A recent study<br />
(He et al. 2004) suggested <strong>the</strong> differences <strong>in</strong> <strong>virulence</strong> was due to <strong>the</strong> presence <strong>of</strong><br />
two pathogencity islands found <strong>in</strong> PA14 that were absent <strong>in</strong> PAO1, named<br />
<strong>Pseudomonas</strong> aerug<strong>in</strong>osa pathogenicity island 1 (PAPI-1) and <strong>Pseudomonas</strong><br />
aerug<strong>in</strong>osa pathogenicity island 2 (PAPI-2).<br />
<strong>The</strong> aim <strong>of</strong> <strong>the</strong> project is to compare <strong>the</strong> contribution <strong>of</strong> PAPI-1 and PAPI-2 to <strong>the</strong><br />
<strong>virulence</strong> <strong>of</strong> P. aerug<strong>in</strong>osa PA14 with<strong>in</strong> a mur<strong>in</strong>e acute pneumonia model.<br />
Importantly, this is <strong>the</strong> first project to look at <strong>the</strong> effects <strong>of</strong> pathogenicity islands on<br />
<strong>in</strong> vivo respiratory <strong>in</strong>fection. To achieve this aim, three isogenic pathogenicity<br />
island deletant mutants were created <strong>in</strong> <strong>the</strong> stra<strong>in</strong> P. aerug<strong>in</strong>osa PA14, a mutant<br />
with PAPI-1 deleted, ano<strong>the</strong>r with PAPI-2 deleted and a third mutant with both<br />
PAPI-1 and PAPI-2 deleted. <strong>The</strong> PA14 mutants and wild-type PA14 were also<br />
compared aga<strong>in</strong>st PAO1. Here<strong>in</strong>, <strong>the</strong> mutants will be referred to as ∆PAPI-1,<br />
∆PAPI-2 and ∆PAPI-1∆PAPI-2.<br />
This results chapter proceeds by describ<strong>in</strong>g <strong>the</strong> mutants used, followed by results<br />
generated whilst develop<strong>in</strong>g <strong>the</strong> mur<strong>in</strong>e <strong>in</strong>fection models, <strong>the</strong> next section describes<br />
<strong>the</strong> results generated us<strong>in</strong>g an acute respiratory model to compare <strong>virulence</strong> and<br />
ends with <strong>the</strong> discussion and conclusions generated from this project.<br />
4.1 Construction <strong>of</strong> <strong>the</strong> deletant mutants and <strong>in</strong> vitro data<br />
Three isogenic pathogenicity island mutants were created by Ewan Harrison<br />
(University <strong>of</strong> Leicester, PhD, unpublished); ∆PAPI-1, ∆PAPI-2 and ∆PAPI-<br />
1∆PAPI-2. <strong>The</strong> first mutants <strong>of</strong> its k<strong>in</strong>d reported <strong>in</strong> P. aerug<strong>in</strong>osa. <strong>The</strong>y were<br />
created us<strong>in</strong>g a mutagenesis system developed by Choi and Schweizer (2005)<br />
specifically target<strong>in</strong>g P. aerug<strong>in</strong>osa. A number <strong>of</strong> <strong>in</strong> vitro test were performed on<br />
PA14 and its isogenic mutants by Ewan Harrison, to assess swarm<strong>in</strong>g mobility,<br />
twitch<strong>in</strong>g mobility, pyocyan<strong>in</strong> production, bi<strong>of</strong>ilm production and growth<br />
characteristics. <strong>The</strong>re were no significant differences reported between PA14 and<br />
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