A Function of SmeDEF, the Major Quinolone Resistance Determinant of Stenotrophomonas maltophilia, Is the Colonization of Plant Roots

Abstract

Quinolones are synthetic antibiotics, and the main cause of resistance to these antimicrobials is mutation of the genes encoding their targets. However, in contrast to the case for other organisms, such mutations have not been found in quinolone-resistant Stenotrophomonas maltophilia isolates, in which overproduction of the SmeDEF efflux pump is a major cause of quinolone resistance. SmeDEF is chromosomally encoded and highly conserved in all studied S. maltophilia strains; it is an ancient element that evolved over millions of years in this species. It thus seems unlikely that its main function would be resistance to quinolones, a family of synthetic antibiotics not present in natural environments until the last few decades. Expression of SmeDEF is tightly controlled by the transcriptional repressor SmeT. Our work shows that plant-produced flavonoids can bind to SmeT, releasing it from smeDEF and smeT operators. Antibiotics extruded by SmeDEF do not impede the binding of SmeT to DNA. The fact that plant-produced flavonoids specifically induce smeDEF expression indicates that they are bona fide effectors regulating expression of this resistance determinant. Expression of efflux pumps is usually downregulated unless their activity is needed. Since smeDEF expression is triggered by plant-produced flavonoids, we reasoned that this efflux pump may have a role in the colonization of plants by S. maltophilia. Our results showed that, indeed, deletion of smeE impairs S. maltophilia colonization of plant roots. Altogether, our results indicate that quinolone resistance is a recent function of SmeDEF and that colonization of plant roots is likely one original function of this efflux pump.

Vitamin K 3 Induces the Expression of the Stenotrophomonas maltophilia SmeVWX Multidrug Efflux Pump

“…For instance, the AcrAB-TolC system in Escherichia coli can be induced by bile salts (29), and in Pseudomonas aeruginosa, the MexXY-OprM system is induced in response to antibiotics that target the ribosome and under oxidative stress conditions (30,31). In the case of S. maltophilia, the SmeDEF efflux pump is induced by triclosan and some plant-produced compounds, which bind its local repressor SmeT, so that smeDEF transcription becomes activated (26,32). To date, it is not known if there is any compound able to induce the expression of the SmeVWX efflux pump; consequently, we carried out a screening of such potential effectors in S. maltophilia PBT02 using compounds belonging to different categories, including antibiotics, compounds that produce oxidative stress, chelating agents, biocides, etc.…”

“…Different compounds from different categories were used in this assay at serial concentrations. Among them were the antibiotics erythromycin (512, 256, and 128 g/ml), gentamicin (32,16, and 8 g/ml), co-trimoxazole (trimethoprimsulfamethoxazole, 1:5) (1, 0.5, and 0.25 g/ml), chloramphenicol (16,8, and 4 g/ml), tobramycin (512, 256, and 128 g/ml), ofloxacin (4, 2, and 1 g/ml), kanamycin (512, 256, and 128 g/ml), tetracycline (4, 2, and 1 g/ml), polymyxin B (2, 1, and 0.5 g/ml), and colistin (24, 12, and 6 g/ml); heavy metals ZnSO 4 (5, 2.5, and 1.25 mM), CuSO 4 (5, 2.5, and 1.25 mM), CdSO 4 (5, 2.5, and 1.25 mM), CoSO 4 (5, 2.5, and 1.25 mM), and FeCl 3 (5, 2.5, and 1.25 mM); oxidative stress compounds paraquat (5, 2.5, and 1.25 mM), vitamin K 3 (2, 1, and 0.5 mM), and tert-butyl hydroperoxide (1, 0.5, and 0.25 mM); biocides triclosan (10, 5, and 2.5 g/ml) and hexachlorophene (100, 50, and 25 g/ml); plant-produced flavonoids phloretin (10, 5, and 2.5 g/ml), quercetin (10, 5, and 2.5 g/ml), and genistein (10, 5, and 2.5 g/ml); detergents SDS (200, 100, and 50 mM) and Tween 20 (200, 100, and 50 mM); chelating agents EDTA (2, 1, and 0.5 mM) and EGTA (2, 1, and 0.5 mM); the analgesic metamizol (10, 5, and 2.5 mg/ml); and the inhibitor of oxidative phosphorylation carbonyl cyanide m-chlorophenyl hydrazone (CCCP; 20, 10, and 5 M). The compounds vitamin K 3 , vitamin K 2 , plumbagin, and tert-butyl hydroperoxide were studied in more detail using a wider range of concentrations.…”

“…In addition, the fact that vitK3 is produced by plants and that S. maltophilia is a common plant inhabitant (54) strongly suggests that SmeVWX might be involved in bacterium-plant interactions in nature, a functional role that has been already demonstrated for the S. maltophilia SmeDEF efflux pump (32).…”

The Challenge of Efflux-Mediated Antibiotic Resistance in Gram-Negative Bacteria

“…Triclosan not only selects SmeDEF-overproducing multidrug-resistant isolates (577) but also induces SmeDEF production via its binding to SmeT and subsequent release from the smeT promoter (865). Similarly, natural flavonoids bind to SmeT and thus influence SmeDEF expression (866). While fusaric acid-inducible FuaABC is positively controlled by its local regulator, FuaR (583), another ABC-type MacABC pump was not experimentally proven to be affected by the MacRS two-component system encoded by genes divergently transcribed from macABC (584).…”

The Efflux Pump SmeDEF Contributes to Trimethoprim-Sulfamethoxazole Resistance in Stenotrophomonas maltophilia

“…Its overexpression, usually due to mutations in the regulator SmeT, reduces its susceptibility to several antibiotics (12), whereas the deletion of smeE makes S. maltophilia more susceptible to such antibiotics (3). Some works have shown that SmeDEF is ubiquitously present in S. maltophilia strains from different origins (13,14) and that despite being a relevant quinolone resistance determinant in this bacterial pathogen, it is involved in S. maltophilia colonization of the roots of plants (15).…”