Use of Non-mammalian Models to Assess the Virulence of Pseudomonas aeruginosa
thesisposted on 2015-08-12, 15:36 authored by Marialuisa Crosatti
Pseudomonas aeruginosa (P. aeruginosa) is the archetypical opportunistic pathogen that accounts for significant numbers of cases of hospital-acquired infections. It adapts to diverse environments in part because it may “modify” its genome through horizontal gene transfer (HGT). Caenorhabditis elegans (C. elegans) and Acanthamoeba spp. (A. castellanii and A. polyphaga) can be employed as in vivo surrogates to investigate the virulence associated with P. aeruginosa clinical isolates and with the presence of genomic islands, genetic elements acquired through HGT. A PA14 based P. aeruginosa pathogenicity island (PAPI)-2 minus mutant exhibited only a minor reduction in virulence as measured by C. elegans survival using the slow-killing assay (SKA) model of virulence but significant attenuation of virulence was found as assessed by the expansion of A. castellanii but not A. polyphaga film on non-nutrient agar. A C. elegans highthroughput assay (HTA) was also established as model of infection and validated against the SKA. The HTA was employed to screen a panel of P. aeruginosa mutants created by deletion of tRNA-associated genomic islands. Few mutants (5 out of 21) showed attenuated virulence toward C. elegans in HTA many of which were confirmed to be less virulent toward the nematode using SKA. Screening of clinical isolates linked the strains isolated from blood culture of a patient with lower survival of C. elegans in HTA compared with strains isolated from sputum. Moreover, type III secretion system effector (T3SS) exoU-positive strains and strains developing pigments during growth were more virulent toward C. elegans in HTA than T3SS effector exoS-positive strains and pigment-less strains, respectively. Since pigment development was also linked to exoU and this gene has been associated with genomic islands integrated at tRNALys10, it was inferred that these genomic islands may bear functions affecting C. elegans survival possibly enhancing iron scavenging.
Date of award2015-08-07
Author affiliationDepartment of Infection, Immunity and Inflammation
Awarding institutionUniversity of Leicester