Involvement of the Putative ATP-Dependent Efflux Proteins PatA and PatB in Fluoroquinolone Resistance of a Multidrug-Resistant Mutant of Streptococcus pneumoniae

Abstract

The multidrug-resistant mutant Streptococcus pneumoniae M22 constitutively overexpresses two genes (patA and patB) that encode proteins homologous to known efflux proteins belonging to the ABC transporter family. It is shown here that PatA and PatB were strongly induced by quinolone antibiotics and distamycin in fluoroquinolone-sensitive strains. PatA was very important for growth of S. pneumoniae, and it could not be disrupted in strain M22. PatB appeared to control metabolic activity, particularly in amino acid biosynthesis, and it may have a pivotal role in coordination of the response to quinolone antibiotics. The induction of PatA and PatB by antibiotics showed a pattern similar to that exhibited by SP1861, a homologue of ABC-type transporters of choline and other osmoprotectants. A second group of quinolone-induced transporter genes comprising SP1587 and SP0287, which are homologues of, respectively, oxalate/formate antiporters and xanthine or uracil permeases belonging to the major facilitator family, showed a different pattern of induction by other antibiotics. There was no evidence for the involvement of PmrA, the putative proton-dependent multidrug transporter that has been implicated in norfloxacin resistance, in the response to quinolone antibiotics in either the resistant mutant or the fluoroquinolone-sensitive strains.Large epidemiological studies of Streptococcus pneumoniae in clinical infections have associated mutations in the genes encoding gyrase and topoisomerase IV with fluoroquinolone resistance (13, 40). However, resistance to fluoroquinolones can also be mediated by active efflux (5, 9-11, 17, 18, 22, 34, 44, 53). Until recently, the only efflux pump implicated in pneumococcal fluoroquinolone resistance was PmrA (22), but it now appears that there must be other efflux pumps involved in this resistance phenotype (11,45). Multidrug-resistant strain M22, a mutant selected after exposure of S. pneumoniae NCTC 7465 (strain M4) to ciprofloxacin, appeared to have such an efflux-mediated resistance mechanism (46). The mutation frequency of 6.9 ϫ 10 Ϫ8 and stable resistance without antibiotic pressure suggested a mutation in a single gene (46). However, no mutations in the fluoroquinolone resistance-determining regions of the A subunits of DNA gyrase or topoisomerase IV have been detected (36). Strain M22 was at least fourfold more resistant than strain M4 to ciprofloxacin, norfloxacin, acriflavine, ethidium bromide, doxorubicin, tetracycline, erythromycin, and cetrimide. The MICs of clinafloxacin, gatifloxacin, grepafloxacin, levofloxacin, and sitafloxacin were reproducibly twofold higher for strain M22 than for strain M4, but those of moxifloxacin, ofloxacin, sparfloxacin, and chloramphenicol were identical for the two strains. The accumulation of ciprofloxacin, gatifloxacin, and ofloxacin by strain M22 was significantly less than that observed in strain M4, whereas the accumulation of norfloxacin and ethidium was consistently higher than in strain M4. Addition of reserpine increased the uptake...

Genome sequencing of linezolid-resistant Streptococcus pneumoniae mutants reveals novel mechanisms of resistance

“…Interestingly, resistance to several other drugs targeting the 23S rRNA was decreased significantly when either spr1887 or spr1885 were inactivated in 1974M2 (Table 3). Consistent with the known role of PatA and PatB in ciprofloxacin resistance (Marrer et al 2006b), inactivation of either of these ABC genes led to a 10-fold decrease in ciprofloxacin resistance back to wild-type levels (Table 3).…”

“…These ABC proteins, also known as PatA and PatB, have been linked to resistance to a number of drugs, including fluoroquinolones (Marrer et al 2006b). This prompted us to test and subsequently verify that both the 1974M1 and 1974M2 mutants, with increased patA and patB expression (Fig.…”

SatR Is a Repressor of Fluoroquinolone Efflux Pump SatAB

“…The presence of an active efflux system was also observed then. We later associated this efflux with the overexpression of SatAB, a norfloxacin and ciprofloxacin ABC transporter showing a high percentage of identity with the fluoroquinolone exporter PatA/PatB of Streptococcus pneumoniae (12,13). The genes encoding this pump form an operon that is preceded by an open reading frame (ORF) encoding a putative MarR superfamily regulator named SatR.…”

Fluoroquinolone Efflux in Streptococcus suis Is Mediated by SatAB and Not by SmrA

“…Several efflux pumps conferring resistance to fluoroquinolones have been described in other Grampositive bacteria. Mainly, members of two families of transporters are involved: (i) proteins belonging to the major facilitator superfamily (MFS), such as Bmr of Bacillus subtilis (29), NorA of Staphylococcus aureus (43), Lde from Listeria monocytogenes (13), and PmrA in Streptococcus pneumoniae (12); and (ii) exporters belonging to the vast ATP binding cassette (ABC) family, such as PatA and PatB from S. pneumoniae (28) or LmrA from Lactococcus lactis (44). Effluxmediated drug resistance is normally a consequence of overexpression of the genes encoding the pumps, either through mutations in regulators or in promoter regions thereof (31).…”