Review Article
J Infect Chemother 1997:3:128-138
Mechanisms of Quinolone Resistance Shinichi Nakamura, PhD Discovery Research Laboratories II, Dainippon Pharmaceutical Co., Ltd., Osaka, Japan
Key words: mechanism, quinolone, resistance, gyrase, topoisomerase IV, permeability
INTRODUCTION Quinolone-resistant organisms are increasing because quinolones are frequently used to c o m b a t a variety of infections. Staphylococcus aureus a n d Pseudomonas aeruginosa are often resistant to quinolones because of c o m m o n quinolone u s e . 1 Quinolone-resistant clinical isolates are also increasing in other organisms such as Echerichia coli, g e n e r a o f Shigella, Serratia a n d Acinetobacter, and also in Neisseria gonorrhoeae though relatively less frequently. 2-4 T h e quinolone resistance genes studied so far are on the bacterial chromosome and are not transmitted by plasmids.5 So quinolone-resistant organisms probably increase through spontaneous m u t a tions in the bacterial chromosome and mutation-bearing strains are selected under circumstances where organisms are in c o n t a c t with a n t i m i c r o b i a l agents including quinolones. Although m a n y quinolone resistance mutations have been reported, the most important for clinical relevance seems to be the mutations of target enzymes such as D N A gyrase (gyrase) and D N A t o p o i s o m e r a s e IV (topo IV), and m u t a t i o n s causing decreased drug accumulation due to decreased drug penetration and/or increased d r u g efflux. T h e m e c h a n i s m s o f q u i n o l o n e resistance have been studied in detail in E. coli, S. aureus and P. aeruginosa.
GYRASE MUTATIONS Gyrase is an enzyme discovered by Gellert et al. in 1976. 6 It is a type II D N A t o p o i s o m e r a s e , which creates a d o u b l e - s t r a n d e d break of D N A for catalyzing D N A supercoiling, relaxation, decatenation, and unknotting. Oyrase has an important role in D N A replication, transcription, and recombination, and is essential to bacterial survival (reviewed in references 7-9). Gyrase holoenzyme of E. coli is a heterotetramer consisting of 2 molecules of each of G y r A and GyrB 1~ which are encoded by the gyrA11 13 (map position: 48 min) and gyrB 14,15 (83 min)
Received Feb. 13, 1997; accepted for publication in revised form, Mar. 6, 1997. *Correspondence and requests for reprints to: Department of Urology, Gifu University School of Medicine, 40 Tsukasamachi, Gifu-shi, Gifu 500, Japan.
128
genes, respectively. G y r A is c o n s i d e r e d a subunit participating in D N A breakage and reunion during enzyme reactions. 7,s G y r B participates in A T P hydrolysis that provides energy for enzyme turnover.16,17 Discovery of the connection between gyrase and quinolone resistance b e g a n with the finding that the nalA (nalidixic acidresistance gene) product is a subunit ofgyrase now called GyrA. is, 19 A year ago, the cou (coumermycin-resistance gene) product was found to be the other subunit of the enzyme, now called GyrB. 2~ F o r about a decade people believed that G y r A alone was responsible for quinolone resistance and GyrB for c o u m a r i n antibiotic resistance. H o w e v e r , cloning a n d c h a r a c t e r i z a t i o n o f the novel nalidixic acid-resistance genes, nalC and nalD, disclosed that they are the m u t a n t alleles ofgyrB. 21 Therefore, it is now thought that quinolone resistance is induced by mutations of b o t h the gyrA and gyrB genes.
gyrA mutations M a n y biochemical studies on gyrA mutations used crude enzyme preparations but the molecular mechanism of quinolone resistance was disclosed after cloning and characterization of the gyrA gene and its mutations.11-13 Quinolone-resistance is induced by a single mutation in the gyrA gene of E. coli. Mutation sites are clustered within a relatively small region at the N - t e r m i n u s of G y r A (amino acid number: 67 to 106), which is called the quinolone resistance determining region (QRDR).22 T h e gyrA mutations cause various levels of resistance to the clinically used quinolones, but mutations causing changes in G y r A serine-83 and/or aspartic acid-87 usually induce high-level resistance to quinolones and are frequently detected in clinical isolates. T h e reason gyrA mutants are c o m m o n in quinolone-resistant clinical isolates seems related to their selective advantage of highlevel resistance to quinolones. W h e n selected with nalidixic acid or enoxacin at relatively low concentrations, the m u t a n t freqencies are almost the same in gyrA and gyrB. 2 3 T h e GyrA m u t a n t with an amino acid change f r o m g l y c i n e - 8 1 to a s p a r t i c a c i d is r e s i s t a n t to fluoroquinolones but unusually susceptible to nalidixic acid. 24 T h e quinolone-resistant m u t a n t gyrA genes are usually recessive to the wild-type allele. 23 T h e quinolone-resistance gyrA mutations have been reported in various organisms such as E. coli, 13'22'24-29
1341-321 X/97/0303-0128/US$3.00 9 JSC/CLJ 1997
Quinolone-resistance mechanisms
Salmonella typhimurium, 3~ S. dysenteriae, 32A. baumanii,33 Aeromonas salmonicida, 34 17. aeruginosa, 35 37 Klebsiella pneumoniae, 38,39a,39b CamFylobacterjejuni 40a C.fetus, 40bN. gonorrhoeae, 41-44 Helicobacter py/or/, 45 Coxiella burnetii, 46 S. aureus, 47-5~ Streptococcus pneumoniae, 52-56 Enterococcus faecalis, 57,58Mycobacterium tuberculosis 59,60 and Mycoplasma hominis 61 (Table 1).The mutations are located within the Q R D R and are identical or similar to those of E. coli in terms of mutation sites and amino acid changes.
gyrB mutations T h e mutations on the gyrB gene are rare in clinical isolates but important for understanding the molecular mechanism of quinolone resistance. Two quinolone-resistance mutations have been reported in g. coli 14 at the center of GyrB which is ~supposed to have the GyrA binding domain.62The mutational change from aspartic acid-426 to asparagine induces resistance to all available quinolones. A mutation from lysine-447 to glutamic acid induces resistance to acidic quinolones such as nalidixic and oxolinic acids but causes hypersusceptibility to amphoteric quinolones with a basic group at position 7. 63 Lysine-447 is supposed to compete with a basic g r o u p of quinolones for ionic interaction with aspartic acid-426. T h e m u t a t i o n from lysine-447 to glutamic acid may facilitate such ionic interaction of quinolones with GyrB and result in the i n d u c t i o n of hypersusceptibility to amphoteric quinolones. 63 T h e quinolone-resistance gyrB genes are also recessive to the wild-type allele. 23 GyrB mutations have been reported in E. coli, 14'63 Neisseria gonorrhoeae 4L64 and S. aureus 48 (Table 2). Similarity in q u i n o l o n e - r e s i s t a n t gyrA and gyrB mutations in various organisms suggests that the mechanism of resistance to quinolones may be similar in all organisms. Quinolones inhibit gyrase action through the f o r m a t i o n o f the t e r n a r y complex o f g y r a s e - D N A quinolone. 65,66 Biochemical studies using enzymes reconstituted from mutant and wild-type subunits indicate that the binding affinity of quinolones to the gyrase-DNA complex is changed by GyrA and/or GyrB mutations. 66 ,67 X-ray crystallographic analysis of a GyrA fragment has revealed that the GyrA mutation sites responsible for q u i n o l o n e resistance are located topologically near tyrosine-122, the site for covalent binding to cleaved DNA, and the possible D N A binding groove of the fragment. 68 TOPOISOMERASE IV MUTATIONS
E. coli topo IV genes were cloned and sequenced by Kato et al. in 1990 as the genes complementing temperaturesensitive par(? and parE (both 65-66 min) mutants which were unable to conduct chromosome partition normally at a high temperature. 69 Topo IV is an enzyme consisting of ParC which has homology with GyrA, and ParE
which has homology with GyrB. 69,70 Topo IV has D N A decatenating and relaxing activities and is thought to play a role in partitioning daughter D N A s at the terminal stage of chromosome replication. 71 Recently it has been suggested that topo IV may also have a role in D N A synthesis. 72 T h e enzyme is essential to bacterial survival. 69 Soon after the discovery of topo IV it was found that quinolones inhibit topo IV activity. 73 The relationship between the inhibition and antibacterial activity remained unknown until recently, because quinolone-resistant topo IV mutants had not been isolated. However, quinoloneresistant strains with topo IV mutations have now been isolated and investigated in some organisms.
parC (grlA) mutations The levels of resistance in quinolone-resistant gyrA mutants ofE. coli depend on an additional mutation in the parC gene suggesting that topo IV may be the secondary target of quinolones. 72,74,75 Quinolone-resistant par(? and wild-type p a r ~ are codominant. In the heterozygote, quinolone susceptibility is midway in quinolone susceptibility b e t w e e n h o m o z y g o u s wild-type and quinolone-resistant strains. 72 This is in contrast with quinolone-resistant gyrA which is recessive to wild-type gyrA +. In N. gonorrhoeae, strains with low-level resistance to quinolones have gyrA mutations alone and strains with high-level resistance contain both gyrA and parC mutations. 43,76a,76b Topo IV genes ofS. aureus were cloned and sequenced as gyrase-like genes, grlA and grlB. These genes have homology with the gyrA and gyrB genes of S. aureus, Bacillus subtilis, and E. coli, and with the E. coli parC and parE genes. 77 S. aureus topo IV is considered the primary target o f quinolones. Clinical isolates with low-level resistance to ciprofloxacin (a quinolone) usually have a grlA mutation alone; isolates with high-level resistance usually possess both grl,4 and gyrA mutations. 77,7s However, this mutation pattern does n o t apply to some quinolones such as sparfloxacin, which seems to act at both topo IV and gyrase equally, 79 and nadifloxacin, whose primary target seems to be gyrase. 8~ The mutant grM gene responsible for quinolone resistance is dominant over the wild-type allele irrespective of gene dosage in transformation with the grlA gene alone, but the dominance depends on gene dosage when transformed with the grlA and grlB genes in combination. 79 The fluoroquinolone resistance gene, flqA, has proved to be the allele of grlA. 8t T h e p r i m a r y target of m o s t quinolones seems to be topo IV52-56,82but in S. pneumoniae sparfloxacin targets gyrase primarily. 83 Thus, whether topo IV or gyrase is the primary target of quinolones depends on both the organisms and the compounds tested. The mutation sites and amino acid changes in ParC (GrlA) have b e e n r e p o r t e d in E. coli, 72,74'75 K. pneumoniae, 3 9 b N. gonorrhoeae, 43 ,7 6 a ,76b S. aureus 77 81 ,84 and
I29
J Infect Chemother 1997:3:128-138 Table 1. Q u i n o l o n e
resistance m u t a t i o n s in the G y r A s u b u n i t o f D N A gyrase.
Organism
Escherichia coli
Amino acid sequence
Reference
67 81 8384 87 106 67- A R V V G DV I G K Y H P H G D S A V Y D T I V RMAQP F S L R Y M L V D G Q - 1 0 6 S CGLP N H DW
V
A
13,22, 24-29
R
G Y
Salmonella typhimurium
30, 31 P
S
F Y
N Y G
Shigella dysenteriae
32 L
Acinetobacter baumanii
.................
E ............. V
Aeromonas salmonicida
. . . . . . . . . . . . . . . . . . . . . . . . . G
Pseudomonas aeruginosa
L .....
33
LP L..D..M
34
........
I
83 87 T .......................
67- . . . . . . . . . . . . . . . .
I
-106
35-37
-106
38,39a, 39b
N
Y G H
Klebsiella pneumoniae
67- . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . F G Y N
Campylobacter jejuni
70-.. T
I..A...R
Campylobacter fetus
71-..
I .............
......
T .... AL ..... I N
D..M..PSIT..-109
T ....
N. .M.VPA
....
N.AM.
I . . . -114
AL .....
40a -110
40b
Y
Nesseria gonorrhoeae
Helicobacter pylori
75- . . I . . . . . . . . . . . . . . . . . . . . . . . . .
71-..
I .............
F
N
Y
G
N ....
AL .....
KV
.V.
D..M.LE
N
.....
41-44
-110
45
-110
46
-114
47-51
V
Y G
Coxiella burnetii
70-.
T ....
L..F
.....
T.C.EAM.L
......
F..PF
....
G 8485
Staphylococcus aureus
I ....
68-.
M ........
SS
88 .EAM .....
LP A
Streptococcus pneumoniae
IT...M
........
D.
.Y.
.P
.....
K G
SS
.EAM .....
WW.Y..M
.....
52-56
K
F Y
Enterococcus faecalis
Mycobacterium tuberculosis
I ....
74-
.
S
.ESM ........
M ..........
.AETM.N
......
R
K
I
G 94
AS C VP
9
.SL ......
Y.P .....
W ....
H
P .....
57, 58
-113
59, 60
N G A Y H
Mycoplasma hominis
..l
....
L . . . . . . . . .
S..E
L
130
AM .....
D...A.P.I
..H
61
Quinolone-resistance mechanisms
S. pneurnoniae5~56,82,83 (Table 3). T h e mutations in ParC (GrlA) c o r r e s p o n d a n d are similar to the q u i n o l o n e resistance mutations in GyrA except for the mutation from alanine- 116 to glutamic acid (or proline) in S. aureus GrlA, and, in S. pneumoniae ParC, the m u t a t i o n from lysine-93 to glutamic acid and the mutation from arginine-95 to cystein. Alanine- 116 of S. aureus GrlA is in the highly conserved region of b o t h GrlA ( P a r C ) and GyrA, and is located in the vicinity of tyrosine-120 of E. coli. The corresponding amino acid ofE. coli GyrA, tyrosine-122, is the site for covalent bonding of a cleaved D N A strand. Lysine-93 and arginine-95 of S. pneumoniae ParC are in the region corresponding to the Q R D R of GyrA.
parE (grlB) mutations It has b e e n reported that the nfxD mutation of E. coli that causes norfloxacin resistance is a mutation in the parE gene. ssa T h e nfxD mutation, inducing an amino acid change from leucine-445 to histidine, causes a minimal inhibitory concentration ( M I C ) increase in a gyrA mutant but not in a gyrA + strain. Leucine-445 corresponds to leucine-451 of GyrB, 4 amino acids downstream of lysine-447 whose mutation causes nalidixic acid resistance.14 T h e pare mutation of S. pneumoniae inducing an amino acid change from aspartic acid-435 to a sparagine also confers low-level resistance to quinolones, sSb T h e m e c h a n i s m of quinolone resistance in topo IV
Table 2. Q u i n o l o n e resistance mutations in the GyrB subunit of D N A gyrase. Organism
Escherichia coli
Neisseria gonorrhoeae
Staphylococcus aureus
A m i n o acid sequence
Reference
426 447 4 2 3 - V E G D S A G G S A K Q G R N R K N Q A I L PLKGK I L N V - 4 5 3 N E 419 416- . . . . . . . . . . M. . .D. . F ............. -436 N 437 458 434- . . . . . . . . . T. S..DSRT ...... R ...... -464 N q
1 4, 63
41, 64
48
The sites and the kinds of a m i n o acids c h a n g e d by q u i n o l o n e resistance mutations are indicated b e l o w the w i l d - t y p e a m i n o acid sequence in each organism. Dots mean the same a m i n o acids as those at the c o r r e s p o n d i n g sites in E. coll. The m a j o r mutation sites are n u m b e r e d a b o v e the w i l d type a m i n o acid sequences. A b b r e v i a t i o n s of a m i n o acids: A, alanine; C, cystein; D, aspartic acid; E, g l u t a m i c acid; F, p h e n y l a l a n i n e ; G, glycine; H, histidine; I, isoleucine; K, lysine; L, leucine; M, met h i o n i n e ; N, asparagine; P, p r o l i n e ; Q, glutamine; R, arginine; S, serine; T, threonine; V, valine; W, t r y p t o p h a n ; and Y, tyrosine.
T a b l e 3. Q u i n o l o n e resistance mutations in the ParC(GrlA) subunit of D N A topoisomerase IV.
Organism
E. coli
A m i n o acid sequence 78 80 84 64-ARTVGDVLGKYHPHGDSACYEAMVLMAQPFSYRYPLVDGQGNWGAPDDPKS FAAMRYTESRL-125 D L K R I
K. pneumoniae
S. aureus
S. pneumoniae
- 1 09
39b
G K
88 91 SA . . . . . R . . . D . T L . . . . I . . I . . F .SR.GDGA-. NIP K G 80 84 64-.K . . . . . I . Q . . . . . . . SV . . . . . RLS .DWKL .HV. I EMH..N. SI . N D P P - . F K Y 79 83 93 95 63-. K S . . N I M . N F . . . . . . SI . D . . . R . S . N W K N . E I . . E M H . . N . SM.GDPP-. Y HT E C F Y 71-..V.. E I ..........
72, 74, 75
G
64-. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . I R
N. gonorrhoeae
Reference
......
116 ...... E P ......
A . . -131
43, 76a, 76b
AK. -124
77-81,84
A..-123
52 56,82,83
131
J Infect Chemother 1997:3:128-138
may be similar to that in gyrase; the sites and the amino acid changes of quinolone-resistant topo IV mutations are similar to those of quinolone-resistant gyrase m u t a tions in m o s t cases. MUTATIONS O N DRUG ACCUMULATION It has been known for years that decrease in permeability to the d r u g is responsible for bacterial resistance to quinolone, s6,s7 Recent studies have disclosed that active efflux of quinolones is also an i m p o r t a n t m e c h a n i s m by which quinolone accumulation is decreased in bacterial cells. In gram-negative organisms, various mutations that resist drug accumulation usually induce changes in outer m e m b r a n e proteins and activate drug-efflux p u m p s concurrently. In gram-positive organisms, mutations that affect drug accumulation activate drug-efflux p u m p s alone (Table 4).
Gram-negativeorganisms In E. coli, mutations that confer nalidixic acid resistance, nalB (58 min) 86,s7 and nalD (89 min), ss decrease nalidixic acid penetration but the m e c h a n i s m remains to be
clarified. T h e mechanisms of resistance of other m u t a tions that resist nalidixic acid, crp, cya, icd, purB, and ctr, are also unknown, s9 After the development of the first new q u i n o l o n e , norfloxacin, 2 norfloxacin-resistant transport mutations norB (34 min) and norC (8 min) were found. 9~T h e norB mutation induces resistance to quinolones and antibiotics, and reduces the a m o u n t of O m p F on the outer m e m b r a n e of bacteria. O m p F is one of the m a j o r porin proteins on the outer m e m b r a n e which serve as channels for the passive diffusion of small hydrophilic molecules including norfloxacin.The norC m u t a t i o n exhibits r e s i s t a n c e to h y d r o p h i l i c quinolones but shows hypersusceptibility to hydrophobic quinolones, hydrophobic antibiotics, dyes and detergents. T h e mutation induces both the reduction of O m p F and an alteration in the lipopolysaccharide structure. Other norfloxacin resistance mutations nfxB (19 min), 91 nfxC (33.7 min), 92 and a ciprofioxacin resistance mutation cfxB (34 min) 93 also reduce O m p F , probably by inhibiting its production through the overexpression of the micF gene that codes for a small R N A c o m p l e m e n t a r y to the 5'-end region of ompFmRNA.94The cfxB gene has been identified as the m u t a n t allele of marR.95 Multiple anti-
Table 4. Quinolone resistance mutations causing decreased drug accumulation. Organism E. coil
P. aeruginosa
P. vulgaris C. jejuni S. aureus S. epidermidis B. subtilis M. smegmatis
Q, quinolone.
132
Gene
Map (min)
Resistance
nalB nalD crp cya icd purB ctr norB norC ~ nfxB nfxC cfxB(marR) marRAB(soxQ) marC(soxR) soxRS soxQ(marA) emrRAB mcb
58 89 74 86 26 25 ? 34 8 19 34 34 34 92 92 34 57.5 --
rob nalB(mexR), cfxB nfxB nfxC pqr ? norA(flqB) norA bmr lfr
? 30 4-8 46
nalidixic acid nalidixic acid nalidixic acid nalidixic acid nalidixic acid nalidixic acid nalidixic acid norfloxacin hydrophilic Qs, norfloxacin norfloxacin multi-drugs multi-drugs multi-drugs multi-drugs multi-drugs multi-drugs microcin B17, sparfloxacin multi-drugs Qs, antibiotics Qs Qs, carbapenems
Mechanism of resistance ?
? ? ? ? ? Om 3F Om ~F $, LPS change Om ~F $, MicF 1` Om :~F $, MicF 1` Om ~F $, MicF 1", Q efflux Om ~F ,[,, MicF 1", Q efflux Om 3F ,[,, MicF 1", Q efflux Om 3F $, MicF 1", Q efflux Om 3F $, MicF 1", Q efflux nalidixic acid efflux 1" sparfloxacin efflux 1"
1` 1", gene activation 1" 1", gene activation 1", gene activation
OmpF $, MicF 1", gene activation OprM 1", Q efflux 1" OprJ 1", Q efflux 1" OprN 1", OprD $, Q efflux 1"
hydrophilic Q efflux 1" hydrophilic Q efflux 1`
Reference 86,87 88 89 89 89 89 89 9O 9O 91 92 93,95 96-101 105 102-104 103 106 107 108 109-115 116-120 121-123 127, 128 129 130-137 138 139 140
Quinolone-resistance mechanisms
biotic resistance mutations (mar) 96 c a u s e resistance to q u i n o l o n e s by a m e c h a n i s m similar to that o f the quinolone resistance genes. 97 Alterations at the marRAB operon (34 min) produce resistance to a wide range of antibacterial agents including quinolones, chloramphenicol, tetracycline, and fl-lactams.98 T h e marR gene codes for the repressor of the mar operon and the marA gene for a positive transcriptional activator. 99,1~176 T h e overproduction of MarA, due to the inactivation of M a r R 95 or a gene dosage effect, 1~ confers the multiple antibiotic resistance p h e n o t y p e . T h e activation of superoxide response genes soxRS (92 rain) by superoxide-generating drugs such as menadione or paraquat induces multiple antibiotic resistance like that of the mar operon.102,103 Such activation represses ompFexpression through the production ofmicFantisense R N A ) ~ One of the sdx genes, soxQ (34 min), seems to be an allele of marA 1~ and one of the mar genes, marC (92 min), seems to be an allele of soxR.105 MarA and SoxS are homologous to each other in amino acid sequence and act on various genes as transcriptional activators) ~ MarA and SoxS likely enhance the expression of a gene coding for a y e t - u n k n o w n quinolone efflux pump. Therefore, the mechanisms of multiple antibiotic resistance i n d u c e d by these genes are probably similar. Mutations in the emrRAB operon (57.5 min) confer resistance to carbonylcyanide mchlorophenylhydrazone and also to a n u m b e r of other toxic compounds including nalidixic acid. 1~ EmrB is homologous to QacA, a p u m p that induces multidrug resistance of S. aureus. 1~ T h e plasmid-encoded mcb operon, which is responsible for the production of an antigyrase protein, microcin B17, and the emrRAB operon share a c o m m o n negative regulator E m r R . T h e M c b E F G pump is dedicated to microcin B17 extrusion but can confer resistance to unrelated c o m p o u n d s such as sparfloxacin, carbonylcyanide m-chlorophenylhydrazone, and tetrachlorosalicylanilide) ~ T h e overexpression of Rob which binds the replication origin, oriC, confers multiple antibiotic resistance similar to that known for the SoxS and M a r A overproduction. 1~ In P. aeruginosa, 3 well-studied quinolone-resistance genes, nalB (30 min), nfxB (4-8 min) and nfxC (46 min) are known.The nalB mutants are resistant to quinolones, tetracycline, chloramphenicol, and erythromycin.l~ The m u t a n t s show d e c r e a s e d q u i n o l o n e a c c u m u l a t i o n which is released by the addition of carbonylcyanide mchlorophenylhydrazone. The cfxB mutation is identical to the nalB mutation) l~ The outer membrane protein OprM increases in nalB mutants.111 O p r M is revealed as the oprKproduct of the mexAB-oprK operon, 112,1x3 and the operon is renamed mexAB-oprM. O p r M is constitutively expressed even in the wild-type strain and is responsible for intrinsic drug resistance of/?. aeruginosa.114 It has been found that nalB is a mutant allele of mexR, a repressor gene for the mexAB-oprM operon. NalB induces coordinate overproduction of MexA, MexB, and OprM which constitute a complex of drug-effiux pump and channel
for d r u g excretion.115 MexR is homologous to M a r R of
E. coli and is supposed to act as not only a repressor but also as an activator.115 T h e nfxB mutants are resistant to q u i n o l o n e s but h y p e r s u s c e p t i b l e to fl-lactams and aminoglycosides, and they show lower norfloxacin uptake. 116 The outer membrane protein OprJ increases in the nfxB mutants)16-118 NfxB is homologous to D N A binding proteins which regulate gene expression.a 19 Cloning of the oprJ and nearby genes has disclosed the mexCD-oprJ operon which is homologous to the mexABoprM operon. 120The nfxB gene is located upstream of the mexCD-oprJ operon just as the nalB gene is located upstream of the mexAB-oprM operon, and nfxB is thought to regulate the o p e r o n ) 2~T h e nfxC mutants are resistant to quinolones, imipenem, and chloramphenicol but hypersusceptible to fl-lactams and aminoglycosides, x21 In nfxC mutants norfloxacin accumulation is decreased as is OprD, while the~amofiht of O p r N shows an increase.a21,122The decrease of O p r D turns out not to be related to quinolone resistance. 123 T h e operon including o w n has been cloned and the gene structure of mexEF-oprN found to be similar to that of mexAB-oprM and mexCD-oprJ) 24 It is postulated that the transcriptional activator gene upstream of the mexEF-oprNoperon corresponds to the nfxC gene. 124 A loss of O p r F in enoxacin-resistant mutants 125 and a quinolone-hypersusceptibility mediated by O p r H overexpression 126 have been reported but details are still unknown. T h e pqr gene in Proteus vulgaris, 127'128 and a yet-unknown gene in Campylobacterjejuni 129have been reported as m u t a t i o n s of p u m p s e x c r e t i n g d r u g s i n c l u d i n g quinolones in gram-negative organisms.
Gram-positive organisms In S. aureus the norfloxacin-resistance gene, norA, has been cloned and sequenced./3~ The product is thought to be a membrane protein acting as a quinolone effiux pump. The overexpression of the nora gene confers resistance to low molecular weight compounds including rodamine 6G, ethidium bromide, relatively hydrophilic quinolones but not relatively hydrophobic quinolones. 131,132However, hydrophilicity-hydrophobicity of quinolones is not an exclusive factor for NorA-mediated quinolone effiux and the bulkiness of the groups at the positions 7 and 8 of a quinolone ring also correlates with the effiux. ~33Quinolone effiux p r o m p t e d by N o r A is inhibited by the addition of carbonylcyanide m-chlorophenylhydrazone which suggests that the effiux is energy-dependent.131 In quinoloneresistant clinical isolates of S. aureus, overexpression of the norA genO 34 and the insertion of the transposon IS256, which has a potent promoter, have been reported.135 The fluoroquinolone resistance mutation, flqB, proves to be a mutation in the promoter region of the norA gene. 136 Multidrug resistance mediated by the nora gene seems to be inducible, x37 Homologs of norA have been reported in S. epidermidis (norA), 138 B. subtilis (bmr) 139 and M. smegmatis (lfr). 14~
133
l Infect Chemother I997:3:128-138
CONCLUSION Q u i n o l o n e r e s i s t a n c e is i n d u c e d b y m u t a t i o n s o n q u i n o l o n e t a r g e t e n z y m e s s u c h as gyrase a n d t o p o IV, a n d b y m u t a t i o n s t h a t p r e v e n t d r u g a c c u m u l a t i o n as a result o f c h a n g e s in o u t e r m e m b r a n e p r o t e i n s a n d / o r activation o f d r u g - e f f i u x p u m p s . M u t a t i o n s o n t h e target e n z y m e s u s u a l l y c a u s e r e s i s t a n c e to q u i n o l o n e s s p e cifically, b u t m u t a t i o n s a f f e c t i n g d r u g a c c u m u l a t i o n c o n f e r r e s i s t a n c e to m u l t i p l e d r u g s . I n m o s t cases a single m u t a t i o n d o e s n o t c a u s e h i g h - l e v e l r e s i s t a n c e to q u i n o l o n e s , b u t m u l t i p l e m u t a t i o n s do. As the f r e q u e n c y o f each m u t a t i o n is a b o u t 10 8, m u l t i p l e m u t a t i o n s h a r d l y o c c u r at t h e s a m e t i m e . T h e r e f o r e , m u t a n t s w i t h h i g h level r e s i s t a n c e are likely to e m e r g e in a stepwise fashion. T h e m o s t i m p o r t a n t clinical p o i n t is t h a t m u t a n t s , even w i t h low levels o f resistance, m u s t n o t b e s e l e c t e d u p o n q u i n o l o n e t r e a t m e n t . I n this c o n t e x t , we m u s t rem e m b e r t h a t s o m e q u i n o l o n e - r e s i s t a n t m u t a n t s m a y be s e l e c t e d n o t o n l y b y q u i n o l o n e derivatives b u t also b y o t h e r k i n d s o f a n t i m i c r o b i a l agents. T h e f u t u r e usefulness o f q u i n o l o n e s as a n t i m i c r o b i a l s m a y d e p e n d o n h o w carefully a n t i m i c r o b i a l agents i n c l u d i n g q u i n o l o n e s are u s e d clinically.
14. 15.
16. 17. 18.
19.
20. 21. 22.
REFERENCES
23. 1. Satake S. Yearly change of drug susceptibility (19831992). In: Medical I n f o r m a t i o n System Developing Center (eds) Drug susceptibility information 1993. Tokyo: Yakugyojihosha, 1993:45-57 (in Japanese). 2. Goldstein FW, Acar JE Epidemiology of quinolone resistance: Europe and North and South America. Drugs 1995;49 (suppl 2) :36-42. 3. Turnidge J. Epidemiology of quinolone resistance: Eastern hemisphere. Drugs 1995;49(suppl 2):43-47. 4. Konno M, Ubukata K. Present status of quinolone resistance. In: UedaY, Shimizu K, Konno M, Matsumoto F (eds) Quinolones.Tokyo: Life Science, 1991:75-84 (in Japanese). 5. Courvalin P. Plasmid-mediated 4-quinolone resistance: a real or apparent absence? Antimicrob Agents Chemother 1990;34:681-684. 6. Gellert M, Mizuuchi K, O ' D e a M H , Nash HA. D N A gyrase: an enzyme that introduces superhelical turns into DNA, Proc Natl Acad Sci U S A 1976;73:3872-3876. 7. Gellert M. D N A topoisomerases. Annu Rev Biochem 1981;50:879-910. 8. Wang JC. D N A topoisomerases. A n n u Rev Biochem 1985;54:665-697. 9. Wang JC. D N A topoisomerases. Annu Rev Biochem 1996;65:635-692. 10. Klevan L, Wang JC. Deoxyribonucleic acid gyrase-deoxyribonucleic acid complex containing 140 base pairs of deoxyribonucleic acid and an a 2 b 2 core. Biochemistry 1980; 19:5229-5234. 11. Swanberg SL, Wang JC. Cloning and sequencing of the Escherichia coligyrA gene coding for the A subunit of D N A gyrase. J Mol Biol 1987;197:723-736. 12. Hussain K, Elliot EJ, Salmond GPC. The ParD- mutant ofEscherichia coli also carries a gyrAam mutation.The complete sequence ofgyrA. Mol Microbiol 1987; 1:259-273. 13. Yoshida H , K o j i m a T, Yamagishi J, N a k a m u r a S.
134
24.
25.
26.
27.
28.
29.
30.
31.
Quinolone-resistant mutations of the gyrA gene of Escherichia coll. Mol Gen Genet 1988;211:1-7. Yamagishi J, Yoshida H,Yamayoshi M, Nakamura S. Nalidixic acid-resistant mutations of the gyrB gene of Escherichia coil Mol Gen Genet 1986;204:367-373. AdachiT, Mizuuchi M, Robinson EA, Appella E, O ' D e a M H , Gellert M, Mizuuchi K. D N A sequence of the E. coli gyrB gene: application of a new sequencing strategy. Nucleic Acids Res 1987; 15:771-784. Sugino A, Cozzarelli NR. The intrinsic ATPase of D N A gyrase. J Biol Chem 1980;255:6299-6306. Maxwell A, Gellert M. Mechanistic aspects of D N A topoisomerases. Adv Protein Chem 1986;38:69-107. Gellert M, Mizuuchi K, O ' D e a M H , ItohT, Tomizawa J. Nalidixic acid resistance: a second genetic character involved in D N A gyrase activity. Proc Natl Acad Sci USA 1977;74:47724776. Sugino A, Peebles CL, Kreuzer KN, CozzareUi NR. Mechanism of action of nalidixic acid: purification of Escherichia coli naL4 gene product and its relationship to D N A gyrase and a novel nicking and closing enzyme. Proc Natl Acad Sci USA 1977;74:4767-4771. Gellert M, O'Dea M H , Itoh T, Tomizawa J. Novobiocin and coumermycin inhibit D N A supercoiling catalyzed by D N A gyrase. Proc N a t Acad Sci USA 1976;73:4474--4478. Yamagishi J,Yoshida H,Yamayoshi M, Nakamura S, Nalidixic acid-resistant mutations of the gyrB gene of Escherichia coll. Mol Gen Genet 1986;204:367-373. Yoshida H, Bogaki M, N a k a m u r a M, N a k a m u r a S. Quinolone resistance- determining region in the D N A gyrase gyrA gene of Escherichia coil Antimicrob Agents Chemother 1990;34:1271-1272. Nakamura S, Nakamura M, KojimaT, Yoshida H. gyrA and gyrB mutations in quinolone-resistant strains of Escherichia coil Antimicrob Agents Chemother 1989;33:254--255. Cambau E, Borden F, Collatz E, Gutmann L. Novel gyrA point mutation in a strain of Escherichia coil resistant to fluoroquinolones but not to nalidixic acid. Antimicrob Agents Chemother 1993;37:1247-1252. Cullen ME, Wyke AW, Kuroda R, Fisher LM. Cloning and characterization of a D N A gyrase A gene from Escherichia coil that confers clinical resistance to 4-quinolones. Antimicrob Agents Chemother 1989;33:886-894. Heisig P, Schedletzky H, Falkenstein-Paul H. Mutations in the gyrA gene of a highly fluoroquinolone-resistant clinical isolate ofEscherichia coli.Antimicrob Agents Chemother 1993;37:696--701. Oram M, Fisher LM. 4-Quinotone resistance mutations in the D N A gyrase of Escherichia coli clinical isolates identified by using the polymerase chain reaction. Antimicrob Agents Chemother 1991;35:387-389. Vila J, Ruiz J, Marco F, Barcelo A, Goni P, Giralt E, De Anta TJ. Association between double mutation in the gyrA gene of ciprofloxacin resistant clinical isolates of Escherichia coli and MICs. Antimicrob Agents Chemother 1994;38: 2477-2479. Truong QC , Van J-CN, Shlaes D, Gutmann L, Moreau NJ, A novel, double mutation in D N A gyrase A of Escherichia coli c o n f e r r i n g resistance to q u i n o l o n e antibiotics. Antimicrob Agents Chemother 1997;41:85-90. Griggs DJ, Gensberg K, Piddock LJV. Mutations in gyrA gene of quinolone- resistant Salmonella serotypes isolated from humans and animals. Antimicrob Agents Chemother 1996;40:1009-1013. Reyna F, Huesca M, Gonzalez V, Fuchs LY. Salmonella t y p h i m u r i u m gyrA m u t a t i o n s a s s o c i a t e d w i t h fluoroquinolone resistance. Antimicrob Agents Chemother 1995;39:1621-1623.
Quinolone-resistance mechanisms
32. Rahman M, Mauff G, Levy J, Couturier M, Pulverer G, Butzler JP. Detection of 4-quinolone resistance mutation in gyrA gene of Shigella dysenteriae type 1 by PCR. Antimicrob Agents Chemother 1994;38:2488-2491. 33. Vila J, Ruiz J, Goni P, Marcos A, De AntaTJ. Mutation in the gyrA gene of quinolone-resistant clinical isolates of Acinetobacter baumannii. Antimicrob Agents Chemother 1995;39:1201-1203. 34. Oppengaard H, Sorum H. gyrA mutations in quinoloneresistant isolates of fish pathogen Aeromonas salmonicida. Antimicrob Agents Chemother 1994;38:2460-2464. 35. HasegawaY, Kobayashi T, Zouga T, Shimazu M, Nishida M. Resistance mutations ofPseudomonas aeruginosa clinical isolates in the presence of fluoroquinolones. J Jpn Assoc Infect Dis 1996;70:123-131 (in Japanese). 36. Kureishi A, Diver JM, Beckthold B, Schollaardt T, Bryan LE. Cloning and nucleotide sequence of Pseudomonas aeruginosa D N A gyrase gyrA gene from strain PAO 1 and quinolone-resistant clinical isolates. Antiriaicrob Agents Chemother 1994;38:1944-1952. 37. Yonezawa M, Takahata M, Matsubara N,WatanabeY, Narita H. D N A gyrase gyrA mutations in quinolone-resistant clinical isolates of Pseudomonas aeruginosa. Antimicrob Agents Chemother 1995;39:1970-1972. 38. Ando S, Katayama T, Hayakawa S, Ishikawa K, Horiba M, Yanaoka M, et al. Quinolone resistance mutations in Klebsiella pneumoniae. Chemotherapy (Tokyo) 1997;45: 670-675 (in Japanese). 39a. Dimri GP, Das HK. Cloning and sequence analysis ofgyr gene o f Klebsiella pneumoniae. N u c l e i c A c i d s Res 1990;18:151-156. 39b.Deguchi T, Fukuoka A, Yasuda M, Nakano M, Ozeki S, Kanematsu E, Nishino Y, Ishihara S, Ban Y, Kawada Y. Alterations in the GyrA subunit of D N A gyrase and the ParC subunit of topoisomerase IV in quinolone-resistant clinical isolates of Klebsiellapneumoniae. Antimicrob Agents Chemother 1997;41:699-701. 40a. WangY, Huang W M , Taylor DE. Cloning and nucleotide sequence of the CamIrylobacterjejuni gyrA gene and characterization of quinolone resistance mutations. Antimicrob Agents Chemother 1993;37:457-463. 40b.Talor DE, Chau AS-S. Cloning and nucleotide sequence of the gyrA gene from Campylobacterfetus subsp, fetus A T C C 2 7 3 7 4 and characterization of ciprofloxacinresistant laboratory and clinical isolates. Anfimicrob Agents Chemother 1997;41:665-671. 41. Belland RJ, Morrison SG, Ison C, H u a n g W M . Neisseria gonorrhoeae acquires mutations in analogous regions of gyrA and parts in fluoroquinolone-resistant isolates. Mol Microbiol 1994;14:371-380. 42. DeguchiT, Yasuda M, Asano M,Tada K~ Iwata H, Komeda H, et al. D N A gyrase mutations in quinolone-resistant clinical isolates ofNeisseria gonorrhoeae. Antimicrob Agents Chemother 1995;39:561-563. 43. DeguchiT, Yasuda M, Nakano M, Tada K, Ozeki S, Ezaki T, et al. Quinolone-resistant Neisseria gonorrhoeae: corrdlation of alterations in the GyrA subunit of D N A gyrase and the ParC subunit of topoisomerase IV with antimic r o b i a l s u s c e p t i b i l i t y profiles. A n t i m i c r o b A g e n t s Chemother 1996;40:1020-1023. 44. Onodera S, Kishimoto K, Kiyota H, Goto H, Igarashi H, Kawahara G, et al. Analysis of resistance mechanism in new quinolone-resistant Neisseria gonorrhoeae, J Jpn Assoc Infect Dis 1995;69:511-516 (in Japanese). 45. Moore RA, Beckthold B,Wong S, Kureishi A, Bryan LE. Nucleotide sequence of the gyrA gene and characterization of ciprofloxacin-resistant mutants of Helicobacterpylori. Antimicrob Agents Chemother 1995;39:107 111.
46. Musso D, Drancourt M, Osscini S, Raoult D. Sequence of quinolone resistance-determining region of gyrA gene for clinical isolates and for an in vitro-selected quinoloneresistant strain of Coxiella burnetii. Antimicrob Agents Chemother 1996;40:870-873. 47. Goswitz JJ, Willard KE, Fasching CE, Peterson LR. Detection of gyrA gene mutations associated with ciprofloxacin resistance in methicillin-resistant Staphylococcus aureus: analysis by polymerase chain reaction and automated direct D N A sequencing. Antimicrob Agents Chemother 1992;36:1166-1169. 48. Ito H, Yoshida H, Bogaki-Shonai M, Niga T, Hattori H, Nakamura S. Quinolone resistance mutations in the D N A gyrase gyrA and gyrB genes of Staphylococcus aureus. Antimicrob Agents Chemother 1994;38:2014-2023. 49. Sreedharan S, Oram M, Jensen B, Peterson LR, Fisher LM. D N A gyrase gyrA mutations in ciprofloxacin-resistant strains of Staphylococcus aureus. Close similarity with quinolone resistance mutations in Escherichia coli. J Bacteriol 1990;172:7260-7262. 50. TakenouchiT, Ishii C~.Sugawara M, TokueY, Ohya S. Incidence of various gyrA mutants in 451 Staphylococcus aureus strains isolated in Japan and their susceptibilities to 10 fluoroquinolones. Antimicrob Agents Chemother 1995;39: 1414-1418. 51. TokueY, Sugano K, Saito D, NodaT, Ohkura H, Shimosato Y, Sekiya T. Detection of novel mutations in the gyrA gene of Staphylococcus aureus by nonradioisotopic single-strand conformation polymorphism analysis and direct D N A sequencing. Antimicrob Agents Chemother 1994;38:428431. 52. TankovicJ, PerichonB, DuvalJ, CourvalinE Contribution of mutations in g3n'A and parC genes to fluoroquinolone resistance of mutants of Streptococcus pneumoniae obtained in vivo and in vitro. Antimicrob Agents Chemother 1996;40: 2505-2510. 53. Munoz R, De La C a m p a AG. ParC subunit of D N A topoisomerase IV of Streptococcuspneumoniae is a primary target of fluoroquinolones and cooperates with D N A gyrase A subunit in forming resistance phenotype. Anfimicrob Agents Chemother 1996;40:2252-2257. 54. Pan X-S, Ambler J, Mehtar S, Fisher LM. Involvement of topoisomerase IV and D N A gyrase as ciprofloxacin targets in Streptococcustmeumoniae. Antimicrob Agents Chemother 1996;40:2321-2326. 55. Gootz J, Zaniewski R, Haskell S, Schmieder B, Tankovic J, G i r a r d D, et al. Activity of the new fluoroquinolone trovafloxacin (CP-99,219) against D N A gyrase and topoisomerase IV mutants of Streptococcus pneumoniae selected in vitro. Antimicrob Agents Chemother 1996;40: 2691-2697. 56. Janoir C, ZellerV, Kitzis M-D, Moreau NJ, G u t m a n n L. High-level fluoroquinolone resistance in Streptococcus pneumoniae requires m u t a t i o n s in parC and gyrA. Antimicrob Agents Chemother 1996;40:2760-2764. 57. KortenV, H u a n g W M , Murray BE. Analysis by P C R and direct D N A sequencing ofgyrA mutations associated with f l u o r o q u i n o l o n e resistance in Enterococcus faecalis. Antimicrob Agents Chemother 1994;38:2091-2094. 58. Tankovic J, Mahjoubi F, Courvalin P, Duval J, Leclercq R. Development of fluoroquinolone resistance in Enterococcus faecalis and role of mutations in the D N A gyrase gyrA gene. Antimicrob Agents Chemother 1996;40:25582561. 59. Alangaden GJ, Manavathu EK, Vakulenko SB, Zvonok NM, Lerner SA. Characterization of fluoroquinolone-resistant mutant strains of Mycobacterium tuberculosis selected in the laboratory and isolated from patients. Antimicrob Agents
135
J Infect Chemother 1997:3:128-138
Chemother 1995;39:1700-1703. 60. Takiff HE, Salazar I, Guerrero C, Philipp W, Huang WM, Kreiswirth B, et al. Cloning and nucleotide sequence of Mycobacterium tuberculosis gyrA and gyrB genes and detection of quinolone resistance mutations. Antimicrob Agents Chemother 1994;38:773-780. 61. Bebear CM, Bore JM, Bebear C, Renaudin J. Characterization ofMycoplasma hominis mutations involved in resistance to fluoroquinolones. Antimicrob Agents Chemother 1997;41:269-273. 62. Cozzarelli NR. D N A gyrase and the supercoiling of DNA. Science 1980;207:953-960. 63. Yoshida H, Bogaki M, N a k a m u r a M, Yamanaka LM, Nakamura S. Quinolone resistance-determining region in the D N A gyrase gyrB gene of Escherichia coli. Antimicrob Agents Chemother 1991 ;35:1647-1650. 64. Stein DC, Danaher RJ, Cook TM. Characterization of a gyrB m u t a t i o n responsible for low-level nalidixic acid resistance in Neisseria gonorrhoeae. Antimicrob Agents Chemother 1991;35:622-626. 65. Shen LL, KohlbrennerWE,Weigl D, Baranowski J. Mechanism of quinolone inhibition of D N A gyrase. J Biol Chem 1989;264:2973-2978. 66. Yoshida H, Nakamura M, Bogaki M, Ito H, Kojima T, Hattori H, Nakamura S. Mechanism of action of quinolones against Escherichia coli D N A gyrase. Antimicrob Agents Chemother 1993;37:839-845. 67. Willmott CJR, Maxwell A. A single point mutation in the D N A gyrase A p r o t e i n greatly r e d u c e s b i n d i n g of fluoroquinolones to the gyrase-DNA complex. Antimicrob Agents Chemother 1993;37:126-127. 68. Morals Cabral JH, Jackson AP, Smith CV, Shikotra N, Maxwell A, Liddington RC. Crystal structure of the breakage-reunion domain of D N A gyrase. Nature 1997; 388:903-906. 69. Kato J, NishimuraY, Imamura R, Niki H, Hiraga S, Suzuki H. New topoisomerase essential for chromosome segregation in E. coli. Cell 1990;63:393-404. 70. Peng H, Marians KJ. Escherichia coli topoisomerase IV.. Purification, characterization, subunit structure, and subunit interactions. J Biol Chem 1993;268:24481-24490. 71. Adams DE, Shekhtman EM, Zechiedrich EL, Schmid MB, Cozzarelli NR. The role of topoisomerase IV in partitioning bacterial replicons and the structure of catenated intermediates in D N A replication. Cell 1992;71:277-288. 72. K h o d u r s k y AB, Z e c h i e d r i c h E L , C o z z a r e l l i N R . Topoisomerase IV is a target of quinolones in Escherichia coli. Proc N a t Acad Sci USA 1995;92:11801-11805. 73. Hoshino K, Kitamura A, Morrissey I, Sato K, Kato J, Ikeda H. C o m p a r i s o n o f i n h i b i t i o n o f Escherichia coti topoisomerase IV by quinolones with D N A gyrase inhibition. Antimicrob Agents Chemother 1994;38:2623-2627. 74. KumagaiY, Kato J, Hoshino K, AkasakaT, Sato K, Ikeda H. Mutants ofEscherichia coli D N A topoisomerase I v p a r C gene. Antimicrob Agents Chemother 1996;40:710-714. 75. Heisig P. Genetic evidence for a role ofparC mutations in development of high-level fiuoroquinolone resistance in Escherichia coli. Antimicrob Agents Chemother 1996;40: 879-885 76a. Belland RJ, Morrison SG, Ison C, H u a n g W M . Neisseria gonorrhoeae acquires mutations in analoqous regions of gyrA and parC in fluoroquinolone-resistant isolates. Mol Microbiol 1994;14:371-380. 76b. Deguchi T, Yasuda M, Nakano M, Ozeki S, EzakiT, Saito I, Kawada Y. Quinolone-resistant Neisseria gonorrhoeae: Correlation of alterations in the GyrA subunit of D N A gyrase and the ParC subunit of topoisomerase IV with antimicrobial susceptibility profiles. Antimicrob Agents
136
Chemother 1996;40:1020-1023. 77. Ferrero L, Cameron B, Manse B, Lagfieaux D, Crouzet J, Famechon A, Blanche F. Cloning and primary structure of Staphylococcus aureus D N A topoisomerase IV: a primary target of fluoroquinolones. Mol Microbiol 1994; 13: 641-653. 78. Ferrero L, Cameron B, Crouzet J. Analysis of gyrA and grlA mutations in stepwise-selected ciprofloxacin-resistant mutants of Staphylococcus aureus. Antimicrob Agents Chemother 1995;39:1554-1558. 79, Yamagishi J, Kojima T, Oyamada Y, Fujimoto K, Hattori H, N a k a m u r a S, Inoue M. Alterations in the D N A topoisomerase IV grlA gene responsible for quinolone resistance in Staphylococcus aureus. Antimicrob Agents Chemother 1996;40:1157-1163. 80. Oizumi N, Kawabata S, Hirano M. Studies on the mechanism of action of nadifloxacin against quinolone-resistant clinical isolates of Staphylococcus aureus. In: Program and Abstracts of the 44th General Meeting of Western Branch of Japan Society of Chemotherapy. Gifu. December 5-6. 1996: (abstr 31) (In Japanese). 81. Ng EY, Trucksis M, Hooper DC. Quinolone resistance mutations in topoisomerase IV: Relationship to the flqA locus and genetic evidence that topoisomerase IV is the primary target and D N A gyrase is the secondary target of fluoroquinolones in Staphylococcus aureus. Antimicrob Agents Chemother 1996;40:1881-1888. 82. Pan X-S, Fisher LM. Cloning and characterization of the parC and p a r e genes of Streptococcus pneumoniae encoding D N A topoisomerase IV: Role in fluoroquinolone resistance. J Bacteriol 1996;178:4060-4069. 83. Pan X-S, Fisher LM. Targeting of D N A gyrase in Streptococcus pneumoniae by sparfioxacin: Selective targeting of gyrase or topoisomerase IV by quinolones. Antimicrob Agents Chemother 1997;41:471-474. 84. Fukuda H, Hori S, Hiramatsu K. The gr/A mutation in norfloxacin-resistant first-step mutants and clinical isolates of Staphylococcus aureus. J Infect Chemother 1996;2:98-101. 85a. Breines D M , Ouabdesselam S, Ng EY, Tankovic J, Shah S, Soussy CJ, Hooper DC. Quinolone resistance locus nfxD of Escherichia coli is a mutant allele of the pare gene encoding a subunit of topoisomerase IV. Antimicrob Agents Chemother 1997;41 : 175-179. 85b. Perichon B, Tankovic J, Courvalin P. Characterization of a mutation in the parE gene that confers fluoroquinolone resistance in Streptococcuspneumoniae. Antimicrob Agents Chemother 1997;41:1166-1167. 86. Hane M., Wood T. Escherichia coli K-12 mutants resistant to nalidixic acid: genetic mapping and dominance studies. J Bacteriol 1969;99:238-241. 87. Bourguignon G J, Levitt M, Sternglanz R. Studies on the mechanism of action of nalidixic acid. Antimicrob Agents Chemother 1973;4:479-486. 88. Hrebenda J, Heleszko H, Brzostek K, Bielecki J. Mutation affecting resistance ofEscherichia coli to nalidixic acid. J Gen Microbiol 1985;131:2285-2292. 89. Hooper DC,Wolfson JS. Mechanism of bacterial resistance to quinolones. In: Hooper DC,Wolfson JS (eds) Quinolone Antimicrob Agents 2nd ed. Washington: American Society for Microbiology, 1993:97-118. 90. Hirai K, Aoyama H, Suzue S, IrikuraT, Iyobe S, Mitsuhashi S. Isolation and characterization of norfloxacin-resistant mutants of Escherichia coli K-12. Antimicrob Agents Chemother 1986;30:248-253. 91. Hooper DC, Wolfson JS, Souza KS, Tung C, M c H u g h GL, Swartz N. Genetic and biochemical characterization of norfloxacin resistance in Escherichia coll. Antimicrob Agents Chemother 1986;29:639-644.
Quinolone-resistance mechanisms
92. Hooper DC,Wolfson JS, Bozza MA, N g EY. Genetics and regulation of outer m e m b r a n e protein expression by quinolone resistance loci nfxB, nfxC, and cfxB. Antimicrob Agents Chemother 1992;36:1151-1154. 93. Hooper DC,Wolfson JS, Souza KS, Ng EY, McHugh GL, Swartz N. Mechanisms of quinolone resistance in Escherichia coli: Characterization of nfxB and cfxB, two mutant resistance loci decreasing norfloxacin accumulation. Antimicrob Agents Chemother 1989;33:283-290. 94. Matsuyama S, Mizushima S. Construction and characterization of a deletion mutant lacking micF, a proposed regulatory gene for O m p F synthesis in Escherichia coll.J Bacteriol 1985; 162:1196-1202. 95. Ariza RR, Cohen SP, Bacb_hawat N, Levy SB, Demple B. Repressor mutations in the marRAB operon that activate oxidative stress genes and multiple antibiotic resistance in Escherichia coil J Bacteriol 1994; 176:143-148. 96. George AM, Levy SB. Amptifiabte resistance to tetracycline, chloramphenicol, and other antibiotics in Escherichia coli: involvement of a non-plasmid-determined efflux of tetracycline. J Bacteriol 1983;155:531-540. 97. Cohen SP, Mcmurry LM, Hooper DC, Wolfson JS, Levy S. Cross-resistance to fluoroquinolones in multiple-antibiotic resistant (Mar) Escherichia coli selected by tetracycline or chloramphenicol: decreased drug accumulation associated with membrane changes in addition to O m p F reduction. Antimicrob Agents Chemother 1989;33:1318-1325. 98. Nikaido H. Prevention of drug access to bacterial targets: permeability barriers and active efflux. Science 1994;264: 382-388. 99. Cohen S, Hachler H, Levy SB. Genetic and functional analysis of the multiple antibiotic resistance (mar) locus in Escherichia coil J Bacteriol 1993; 175:1484-1492. 100. Jair K-W, Martin RG, Rosner J-L, Fujita N, Ishihama A, WorfJr RE. Purification and regulatory properties of MarA protein, a transcriptional activator ofEscherichia colimultiple antibiotic and superoxide resistance promoters. J Bacteriol 1995;177:7100-7104. 101. Gambino L, Gracheck SJ, Miller PF. Overexpression of the MarA positive regulator is sufficient to confer multiple antibiotic resistance in Escherichia coli. J Bacteriol 1993;175:2888-2894. 102. Greenberg JT, Monach P, Chou JH, Josephy PD, Demple B. Positive control of a global antioxidant defence regulon activated by superoxide-generating agents in Escherichia coli. Proc Natl Acad Sci U S A 1990;87:6181-6185. 103. Greenberg JT, Chou JH, Monach PA, Demple B. Activation of oxidative stress genes by mutations at the soxQ/ cfxB/marA locus of Escherichia coli. J Bacteriol 1991; 173: 4433-4439. 104. Chou JH, Greenberg JT, Demple B. Posttranscriptional repression of Escherichia coli O m p F protein in response to redox stress: positive control of the micFantisense R N A by the soxRS locus. J Bacteriol 1993; 175:1026-1031. 105. Miller PF, Gambino LF, Sulavik MC, Gracheck SJ. Genetic relationship between soxRS and mar loci in promoting multiple antibiotic resistance in Escherichia coil Antimicrob Agents Chemother 1994;38:1773 1779. 106. Lomovskaya O, Lewis K. emr, an Escherichia coli locus for multidrug resistance. Proc Natl Acad Sci USA 1992; 89:8938-8942. 107. Lomovskaya O, Kawai F, Matin A. Differential regulation of the mcb and emr operons of Escherichia coli: role of mcb in multidrug resistance. Antimicrob Agents Chemother 1996;40:1050-1052. 108. Ariza RR, Li Z, Ringstad N, Demple B. Activation of multiple antibiotic resistance and binding of stressinducible promoters by Escherichia coli Rob protein. J
Bacteriol 1995;177:1655-1661. 109. Rella M, Haas D. Resistance of Pseudomonas aeruginosa PAO to nalidixic acid and low levels of ]3-1actam antibiotics: mapping of chromosomal genes. Antimicrob Agents Chemother 1982;22:242-249. 110. Robillard NJ, Scarpa AL. Genetic and physiological characterization of ciprofloxacin resistance in Pseudomonas aeruginosa PAO. Antimicrob Agents Chemother 1988;32: 535-539. 111. Masuda N, Ohya S. Cross-resistance to meropenem, cephems, and quinolones in Pseudomonas aeruginosa. Antimicrob Agents Chemother 1992;36:1847-1851. 112. Gotoh N, Tsujimoto H., Poole K, Yamagishi J, Nishino T. T h e outer membrane protein O p r M of Pseudomonas aeruginosa is encoded by oprK of the mexA-mexB-oprK multidrug resistance operon. Antimicrob Agents Chemother 1995;39:2567-2569. 113. Hamzehpour M M , Pechere J, Plesiat P, KohlerT. OprK and O p r M define two genetically distinct multidrug efflux systems in Pseudomonas aeruginosa. Antimicrob Agents Chemother~ 9957~0:2392-2396. 114. Li X-Z, Livermore DM, Nikaido H. Role ofeffiux pump(s) in intrinsic resistance ofPseudomonas aeruginosa: resistance to tetracycline, c h l o r a m p h e n i c o l , and norfloxacin. Antimicrob Agents Chemother 1994;38:1732-1741. 115. Poole K, Tetro K, Zhao Q, Neshat S, Heinrichs E, Bianco N. Expression of the multidrug resistance operon mexAmexB-oprM in Pseudomonas aeruginosa: mexR encodes a regulator of operon expression. A n t i m i c r o b Agents Chemother 1996;40:2021-2028. 116. Hirai K, Suzue S, IrikuraT, Iyobe S, Mitsuhashi S. Mutations producing resistance to norfloxacin in Pseudomonas aeruginosa. Antimicrob Agents Chemother 1987;31:582586. 117. Hosaka M, Gotoh N, Nishino T. Purification of a 54kilodalton protein (OprJ) produced in NfxB mutants of Pseudomonas auruginosa and production of a monoclonal antibody specific to OprJ. Antimicrob Agents Chemother 1995;39:1731-1735. 118. Masuda N, Gotoh N, Ohya S, Nishino T. Quantitative correlation between susceptibility and OprJ production in NfxB mutants of Pseudomonas aeruginosa. Antimicrob Agents Chemother 1996;40:909-913. 119. OkazakiT, Hirai K. Cloning and nucleotide sequence of the Pseudomonas aeruginosa nfxB gene, conferring resistance to new quinolones. F E M S Microbiol Lett 1992;97: 197-202. 120. Poole K, Gotoh N, Tsujimoto H, Zhao Q, Wasa A,Yamasaki T~ et al. Overexpression of the mexC-rnexD-oprJ ettlux operon in nfxB-type multidrug-resistant strains of Pseudomonas aeruginosa. Mol Microbiol 1996;21:713-724. 121. Fukuda H, Hosaka M, Hirai K, Iyobe S. New norfloxacin resistance gene in Pseudomonas aeruginosa PAO. Antimicrob Agents Chemother 1990;34:1757-1761. 122. Fukuda H, Hosaka M, Iyobe S, Gotoh N, NishinoT, Hirai K. nfxC-type quinolone resistance in a clinical isolate of Pseudomonas aeruginosa. Antimicrob Agents Chemother 1995;39:790-792. 123. H u a n g H, Hancock REW. Genetic definition of the substrate selectivityof outer membrane porin protein OprD of Pseudomonas aeruginosa. J Bacteriol 1993;175:77937800. 124. Kohler T, Michea-Hamzehpour M, Henze U, Gotoh N, Curty LK, Pechere J-C. Characterization of MexE-MexFOprN, a positively regulated multidrug efflux system of Pseudomonas aeruginosa. Mol Microbio11997;23:345-354. 125. Piddock LJV, BeUido F, Bains M, Hancock REW- A pleiotropic, posttherapy, enoxacin-resistant mutant of Pseu-
137
J Infect Chemother I997:3:128-138
126.
127. 128.
129.
130.
131.
132.
133.
138
domonas aeruginosa. Antimicrob Agents C h e m o t h e r 1992;36:1057-1061. Young M, Hancock REW. Fluoroquinolone supersusceptibility m e d i a t e d by outer m e m b r a n e p r o t e i n O p r H overexpression in Pseudomonas aeruginosa: evidence for involvement of a nonporin pathway. Antimicrob Agents Chemother 1992;36:2365-2369. Ishida H, Fuziwara H, KaiboriY, HoriuchiT, Sato K, Osada Y. Cloning of multidrug resistance gene pqrA from Proteus vulgaris. Antimicrob Agents Chemother 1995;39:453-457. Ishii H, Sato K, Hoshino K, Sato M,Yamaguchi A, Sawai T, O s a d a Y. Active efflux of ofloxacin by a highly quinolone-resistant strain of Proteus vulgaris. J Antimicrob Chemother 1991:28:827-836. Charvalos E, Tselentis Y, Hamzehpour M M , Kohler T, Pechere J-C. Evidence for an ettlux pump in multidrugr e s i s t a n t Campylobacter jejuni. A n t i m i c r o b A g e n t s Chemother 1995;39:2019-2022. Ubukata K, Itoh N-Y, Konno M. Cloning and expression of the norA gene for fluoroquinolone resistance in Staphylococcus aureus. Antimicrob Agents Chemother 1989;33:1535-1539. Yoshida H, Bogaki M, Nakamura S, Ubukata K, Konno M. Nucleotide sequence and characterization of the Staphylococcus aureus nora gene, which confers resistance to quinolones. J Bacteriol 1990; 172:6942-6949. Neyfakh AA, Borsch CM, Kaatz GW. Fluoroquinolone resistance protein NorA of Staphylococcus aureus is a multidrug efflux transporter. Antimicrob Agents Chemother 1993;37:128-129. Takenouchi T, Tabata F, Iwata Y, Hanzawa H, Sugawara
134. 135.
136.
137. 138. 139.
140.
M, Ohya S. Hydrophilicity of quinolones is not an exclusive factor for decreased activity in effhix-mediated resistant mutants of Staphylococcus aureus. Antimicrob Agents Chemother 1996;40:1835-1842. K a a t z GW, Seo S M , R u b l e CA. E f f l u x - m e d i a t e d fluoroquinolone resistance in Staphylococcus aureus. Antimicrob Agents Chemother 1993;37:1086-1094. Kojima T, Yamagishi J, OyamadaY, Yoshida H, Hattori H, Inoue M, Nakamura S. Analysis of quinolone resistance genes in a clinical isolate of quinolone-resistant MRSA. Drugs 1995;49(Suppl.2): 182-184. Ng EYW, Trucksis M, Hooper DC. Quinolone resistance mediated by norA: physiologic characterization and relationship toflqB, a quinolone resistance locus on the Staphylococcus aureus chromosome. Antimicrob Agents Chemother 1994;38:1345-1355. Kaatz GW, Seo SM. Inducible NorA-mediated multidrug resistance in Staphylococcus aureus. Antimicrob Agents Chemother 1995;39:2650-2655. Ubukata K. The norA gene conferring new quinolone resistance in Staphylococcus epidermidis. Chemotherapy 1991;39:1001-1013 (in Japanese). Neyfakh AA. The multidrug efflux transporter of Bacillus subtilis is a structural and functional homolog of the Staphylococcus NorA protein. Antimicrob Agents Chemother 1992;36:484-485. TakiffHE, Cimino M, Musso MC,WeisbrodT, Martinez R, Delgado MB, et al. Eftlux pump of the proton antiporter family confers low-level fluoroquinolone resistance in Mycobacterium smegmatis. Proc Natl Acad Sci U S A 1996;93:362-366.
