Abstract
Multidrug efflux pumps constitute a category of antibiotic resistance determinants that are a part of the core bacterial genomes. Given their conservation, it is conceivable that they present functions beyond the extrusion of antibiotics currently used for therapy. Pseudomonas aeruginosa stands as a relevant respiratory pathogen, with a high prevalence at hospitals and in cystic fibrosis patients. Part of its success relies on its low susceptibility to antibiotics and on the production of virulence factors, whose expression is regulated in several cases by quorum sensing (QS). We found that overexpression of the MexCD-OprJ multidrug efflux pump shuts down the P. aeruginosa QS response. Our results support that MexCD-OprJ extrudes kynurenine, a precursor of the alkyl-quinolone signal (AQS) molecules. Anthranilate and octanoate, also AQS precursors, do not seem to be extruded by MexCD-OprJ. Kynurenine extrusion is not sufficient to reduce the QS response in a mutant overexpressing this efflux pump. Impaired QS response is mainly due to the extrusion of 4-hydroxy-2-heptylquinoline (HHQ), the precursor of the Pseudomonas Quinolone Signal (PQS), leading to low PQS intracellular levels and reduced production of QS signal molecules. As the consequence, the expression of QS-regulated genes is impaired and the production of QS-regulated virulence factors strongly decreases in a MexCD-OprN P. aeruginosa overexpressing mutant. Previous work showed that MexEF-OprJ, another P. aeruginosa efflux pump, is also able of extruding kynurenine and HHQ. However, opposite to our findings, the QS defect in a MexEF-OprN overproducer is due to kynurenine extrusion. These results indicate that, although efflux pumps can share some substrates, the affinity for each of them can be different. Although the QS response is triggered by population density, information on additional elements able of modulating such response is still scarce. This is particularly important in the case of P. aeruginosa lung chronic infections, a situation in which QS-defective mutants are accumulated. If MexCD-OprJ overexpression alleviates the cost associated to triggering the QS response when un-needed, it could be possible that MexCD-OprJ antibiotic resistant overproducer strains might be selected even in the absence of antibiotic selective pressure, acting as antibiotic resistant cheaters in heterogeneous P. aeruginosa populations.
Highlights
Pseudomonas aeruginosa is a free-living microorganism able to survive in different environments that plays an ecological role in natural ecosystems (Green et al, 1974; Romling et al, 1994; Morales et al, 2004; Walker et al, 2004), but it is an important causative agent of infections in patients with underlying diseases (Obritsch et al, 2005; Driscoll et al, 2007; Martinez-Solano et al, 2008; Talwalkar and Murray, 2016)
We demonstrate that a P. aeruginosa nfxB∗ mutant, which overexpresses the MexCD-OprJ efflux pump, exhibits an impaired quorum sensing (QS) response due the extrusion of HHQ
This non-physiological extrusion leads to a decrease in the expression of the pqsABCDE operon responsible for AQs synthesis, which affects AQs-dependent and the PqsE-dependent regulons that comprise the genes involved in swarming motility, and in the production of pyocyanin, rhamnolipids, and proteases among others (Hazan et al, 2010; Rampioni et al, 2010, 2016)
Summary
Pseudomonas aeruginosa is a free-living microorganism able to survive in different environments that plays an ecological role in natural ecosystems (Green et al, 1974; Romling et al, 1994; Morales et al, 2004; Walker et al, 2004), but it is an important causative agent of infections in patients with underlying diseases (Obritsch et al, 2005; Driscoll et al, 2007; Martinez-Solano et al, 2008; Talwalkar and Murray, 2016). Relevant is the activity of chromosomally-encoded multidrug resistance (MDR) efflux pumps (Vila and Martínez, 2008; Hernando-Amado et al, 2016). The acquisition of mutation-driven resistance is common in this opportunistic pathogen, along chronic infections (Oliver et al, 2015; Palmer and Whiteley, 2015; Lopez-Causape et al, 2018), where the constitutive overexpression of MDR efflux pumps is one of the biggest problems to eradicate these infections (Vila and Martínez, 2008; Vila et al, 2011; Hernando-Amado et al, 2016). Efflux pumps exhibit different functions, with physiological and ecological significances that go beyond their activity as antibiotic resistance elements (Piddock, 2006; Martinez et al, 2009; Alcalde-Rico et al, 2016; Hernando-Amado et al, 2016). In the case of P. aeruginosa, an opportunistic pathogen not fully adapted to human hosts (Alonso et al, 1999; Morales et al, 2004), these functions should be of relevance for the success of P. aeruginosa as a respiratory infectious pathogen
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