Date Published: September 28, 2018
Publisher: Public Library of Science
Author(s): Dinesh Subedi, Ajay Kumar Vijay, Gurjeet Singh Kohli, Scott A. Rice, Mark Willcox, Abdelwahab Omri.
Virulent strains of Pseudomonas aeruginosa are often associated with an acquired cytotoxic protein, exoenzyme U (ExoU) that rapidly destroys the cell membranes of host cells by its phospholipase activity. Strains possessing the exoU gene are predominant in eye infections and are more resistant to antibiotics. Thus, it is essential to understand treatment options for these strains. Here, we have investigated the resistance profiles and genes associated with resistance for fluoroquinolone and beta-lactams. A total of 22 strains of P. aeruginosa from anterior eye infections, microbial keratitis (MK), and the lungs of cystic fibrosis (CF) patients were used. Based on whole genome sequencing, the prevalence of the exoU gene was 61.5% in MK isolates whereas none of the CF isolates possessed this gene. Overall, higher antibiotic resistance was observed in the isolates possessing exoU. Of the exoU strains, all except one were resistant to fluoroquinolones, 100% were resistant to beta-lactams. 75% had mutations in quinolone resistance determining regions (T81I gyrA and/or S87L parC) which correlated with fluoroquinolone resistance. In addition, exoU strains had mutations at K76Q, A110T, and V126E in ampC, Q155I and V356I in ampR and E114A, G283E, and M288R in mexR genes that are associated with higher beta-lactamase and efflux pump activities. In contrast, such mutations were not observed in the strains lacking exoU. The expression of the ampC gene increased by up to nine-fold in all eight exoU strains and the ampR was upregulated in seven exoU strains compared to PAO1. The expression of mexR gene was 1.4 to 3.6 fold lower in 75% of exoU strains. This study highlights the association between virulence traits and antibiotic resistance in pathogenic P. aeruginosa.
Pseudomonas aeruginosa infections can be severe in people with a compromised immune system and impaired anatomical structures caused by, for example burns, cystic fibrosis or mechanical abrasions . P. aeruginosa is a successful opportunistic pathogen in part due to its production of a diverse repertoire of pathogenic factors and its innate ability to evade the host immune system . Treatment of P. aeruginosa infections can be challenging due to the inherent antibiotic resistance, where some studies have shown that half of the isolates from clinical infections were resistant to antibiotics . Furthermore, reports on co-selection of antibiotic resistance and pathogenic factors indicate that antibiotic resistance may be a factor for the evolution of more virulent strains of P. aeruginosa or vice versa [4–13].
The exoU gene is commonly found in P. aeruginosa strains isolated from contact lens-related microbial keratitis, at frequencies of 46–54%,  whereas it only occurs in 0–14% of non-ocular isolates [6, 22–24]. Similar to a previous report,  exoU+ strains in the current study had higher resistance to beta-lactams than exoS+ strains (100% exoU strains vs. 61% exoS strains were resistant to at least one beta-lactam). ExoU secreting P. aeruginosa had more mutations in genes that are associated with beta-lactam resistance (mexR, ampC and ampR) than did exoS+ strains. Gene expression analysis suggested that such mutations generally lead to antibiotic resistance, as the expression of ampC and ampR generally increased while the expression of mexR was decreased, compared to the sensitive strain PAO1.