Phylogrouping of Pseudomonas aeruginosa from burn patients reveals distinct group 3 characteristics

<b>Introduction.</b> <i>Pseudomonas aeruginosa</i> is a bacterial pathogen that causes acute infections in burned patients which can be very difficult to treat due to the high frequency of antibiotic-resistant strains. In 2019, whole-genome analysis led to the identification of five <i>P. aeruginosa</i> phylogroups including both clinical and environmental isolates. One of these phylogroups (Group 3) is highly genetically divergent and has been recently reclassified as <i>Pseudomonas paraeruginosa</i> based on genomic analysis, although there is no phenotypic difference with other <i>P. aeruginosa</i> phylogroups.<b>Hypothesis.</b> We propose that by using phylogroup-specific sequences, a multiplex PCR could be devised to detect Group 3 strains among clinical isolates from burned patients.<b>Aim.</b> To design a multiplex PCR that can be used to classify strains belonging to phylogroups 1, 2, 3 or 5. It is used to study a collection of 136 <i>P</i>. <i>aeruginosa</i> isolates obtained from 136 burned patients treated in a tertiary hospital in Mexico City during a 10-year period (2011 to 2021) to detect strains belonging to Group 3.<b>Methodology.</b> A multiplex PCR was devised to classify into four of the five <i>P. aeruginosa</i> phylogroups a collection of 136 strains isolated from burned patients. The genome sequence of the three strains belonging to Group 3 or <i>P. paraeruginosa</i> (P701, P1543 and P1565) was analysed bioinformatically, and their production of virulence factors was determined using reported microbiological procedures. Mutant complementation by genes carried in plasmids was performed to characterize the P1543-defective LasR protein. The MIC for 12 antibiotics of the 3 identified Group 3 strains was determined using standard techniques.<b>Results.</b> Using this multiplex PCR, we identified three strains belonging to the PA7 group (2%), which were sequenced and phenotypically characterized. We determined their production of virulence factors such as elastase, motility, biofilm formation, pyocyanin, rhamnolipids and their antibiotic resistance profile and showed that one of these strains (P1543) harbours a point mutation that inactivates <i>lasR</i> which causes elastase deficiency and lack of swarming motility and biofilm formation.<b>Conclusion.</b> These results show that <i>P. aeruginosa</i> Group 3 strains, or <i>P. paraeruginosa</i> isolates, have the same physiopathology as the most common <i>P. aeruginosa</i> phylogroup.