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.

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C. difficile is GI pathogen which under certain conditions colonises the colon leading CDI. During passage through the GI tract the bacterium in either vegetative or spore form encounters innate antimicrobial defence mechanisms evolved to prevent colonisation by microorganisms. One of these natural defences is the production of a potent digestive secretion known as bile. The major role of bile is to emulsify fats from ingested material to aid digestion. However, it is also an effective antimicrobial. The tolerance of C. difficile and its interaction with bile has not yet been fully investigated. The results obtained here describe the tolerance levels of two pathogenic strains, 630△erm and R20291, to bovine and ovine bile. Significant differences were seen in the tolerance of R20291 compared to 630△erm, with R20291 showing tolerance of a much higher concentration of bile. Analysis of genome-wide transcription levels using a microarray revealed differences in the expression of genes in 630△erm in response to bile exposure, although these were not statistically significant. These preliminary investigations do however provide a foundation for further elucidation of the interaction of C. difficile with bile.

Photodynamic therapy is a promising new strategy for the treatment of superficial skin infections and periodontitis. A limitation of antibiotic treatment for these diseases is that even after successful killing of the infecting organism, secreted virulence factors may still be present and cause significant damage to host tissues. If light-activated antimicrobial agents can inactivate microbial virulence factors in addition to killing the pathogenic microorganisms, this would represent an advantage of photodynamic therapy over conventional treatment options. The light-activated antimicrobial agents methylene blue and tin chlorin e6 in combination with laser light of 665 and 633 nm respectively, were assessed for their antibacterial activity and ability to reduce the activity of selected virulence factors of Staphylococcus aureus and Porphyromonas gingivalis. In addition to successfully reducing the microbial burden, it was demonstrated that photosensitisation was able to cause significant reductions in the activity of a number of secreted and cell wall-associated virulence factors produced by these species when irradiated with laser light of the appropriate wavelength. Photosensitisation was also shown to reduce the biological activities of the proinflammatory cytokines tumour necrosis factor-alpha and interleukin-6, which are produced in response to infecting bacteria and are associated with damage to host tissues. The results of these studies indicate that light-activated antimicrobial agents may be useful in reducing the pathology associated with bacterial virulence factors and host-mediated inflammation when used as part of an antimicrobial treatment regimen.

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