Profiling the susceptibility of Pseudomonas aeruginosa strains from acute and chronic infections to cell-wall-targeting immune proteins

Abstract

In the current scenario of high antibiotic resistance, the search for therapeutic options against Pseudomonas aeruginosa must be approached from different perspectives: cell-wall biology as source of bacterial weak points and our immune system as source of weapons. Our recent study suggests that once the permeability barrier has been overcome, the activity of our cell-wall-targeting immune proteins is notably enhanced, more in mutants with impaired peptidoglycan recycling. The present work aims at analyzing the activity of these proteins [lysozyme and Peptidoglycan-Recognition-Proteins (PGLYRPs)], alone or with a permeabilizer (subinhibitory colistin) in clinical strains, along with other features related to the cell-wall. We compared the most relevant and complementary scenarios: acute (bacteremia) and chronic infections [early/late isolates from lungs of cystic fibrosis (CF) patients]. Although a low activity of lysozyme/PGLYRPs per se (except punctual highly susceptible strains) was found, the colistin addition significantly increased their activity regardless of the strains’ colistin resistance levels. Our results show increased susceptibility in late CF isolates, suggesting that CF adaptation renders P. aeruginosa more vulnerable to proteins targeting the cell-wall. Thus, our work suggests that attacking some P. aeruginosa cell-wall biology-related elements to increase the activity of our innate weapons could be a promising therapeutic strategy.

Comparative Analysis of Peptidoglycans From Pseudomonas aeruginosa Isolates Recovered From Chronic and Acute Infections

“…Although not enough to draw any trend among collections, the existence of an appreciable degree of intra-collections variability in the HPLC-derived parameters, made us think about the possibility of linking certain of the structural/compositional features with previously published phenotypes from the same strains (Cabot et al, 2011; López-Causapé et al, 2013; Torrens et al, 2019). In this regard, we sought to see if the phenotypes of susceptibility to different types of aggressions against the peptidoglycan, i.e., ceftazidime MIC and bacterial survival to the treatments of lysozyme + colistin, PGLYRP1 + colistin or PGLYRP2 + colistin (Torrens et al, 2019) (Supplementary Data Set S1), could be correlated with any of the HPLC-derived parameters. The search for correlations by means of the Spearman’s coefficients only provided a statistically significant inverse correlation between the survival to PGLYRP2 + colistin treatment and the relative abundance of stem pentapeptides (Spearman rank correlation coefficient −0.394; p -value 0.014).…”

“…Besides the reference strains ( P. aeruginosa PAO1 and PA14 and Escherichia coli XL-1 blue), the previously described mutant of P. aeruginosa PAOΔdacBΔdacCΔpbpG (Ropy et al, 2015) was used for comparative purposes and as a control for our procedures’ capacity to detect differences in the muropeptides composition. The clinical strains used in this work belong to two previously described collections of clinical isolates, recently analyzed in another work of our group and displayed in the Supplementary Data Set S1 (Torrens et al, 2019). First, twelve strains isolated from bacteremia patients and belonging to a Spanish multicenter study (Cabot et al, 2011) were included.…”

“…Second, we used twelve pairs of CF sequential isogenic isolates [early-late isolate, obtained with a difference of at least 3 years from a total of 10 patients (thus, from two of the patients, two pairs of isolates were obtained on each, at different time points)], proceeding from a previously published work (López-Causapé et al, 2013). The previously determined values of ceftazidime Minimum Inhibitory Concentration (MIC, mg/L), and the percentages of bacterial survival after treatments with: (i) lysozyme (25 mg/L) + colistin (0.025 mg/L) (ii) Peptidoglycan Recognition Protein 1, PGLYRP1 (50 mg/L) + colistin (0.025 mg/L) and (iii) PGLYRP2 (50 mg/L) + colistin (0.025 mg/L) are also displayed in the Supplementary Data Set S1 (Cabot et al, 2011; López-Causapé et al, 2013; Torrens et al, 2019).…”

“…To assess the potential correlation between HPLC-derived parameters and phenotypes of susceptibility/resistance to aggressions against peptidoglycan [ceftazidime MICs and percentages of bacterial survival after treatments with lysozyme/PGLYRP1/PGLYRP2 + colistin (Torrens et al, 2019)], the bivariate associations were evaluated by Spearman’s correlation coefficients. The indicated variables were dichotomized using the median and the comparisons between groups were analyzed using the Mann–Whitney U test.…”

“…Nevertheless, although it is unanimously accepted that the virulence attenuation is a hallmark of P. aeruginosa chronic strains (Cullen and McClean, 2015; Faure et al, 2018), the reduction in the inflammatory capacity is far from being a uniform feature of them. In fact, several works display contradictory results in this regard (Cigana et al, 2009; Hawdon et al, 2010; Di Lorenzo et al, 2015; LaFayette et al, 2015; Torrens et al, 2019). As well, the active search for peptidoglycans with reduced or increased inflammatory capacity has never been conducted in large numbers of P. aeruginosa clinical isolates, but only in punctual strains (Cigana et al, 2009).…”