scholarly journals Novel Ser79Leu and Ser81Ile Substitutions in the Quinolone Resistance-Determining Regions of ParC Topoisomerase IV and GyrA DNA Gyrase Subunits from Recent Fluoroquinolone-Resistant Streptococcus pneumoniae Clinical Isolates

2005 ◽  
Vol 49 (6) ◽  
pp. 2479-2486 ◽  
Author(s):  
Nataliya Korzheva ◽  
Todd A. Davies ◽  
Raul Goldschmidt
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Amino Acid Substitutions ◽  
Topoisomerase Iv ◽  
Isogenic Strains ◽  
New Mutations

ABSTRACT Resistance of Streptococcus pneumoniae to fluoroquinolones is caused predominantly by amino acid substitutions at positions Ser79 of ParC and Ser81 of GyrA to either Phe or Tyr encoded in the quinolone resistance-determining regions of the parC topoisomerase IV and gyrA DNA gyrase genes. Analysis of highly resistant clinical isolates identified novel second-step substitutions, Ser79Leu (ParC) and Ser81Ile (GyrA). To determine contributions of these new mutations to fluoroquinolone resistance either alone or in combination with other Ser79/81 alleles, the substitutions Ser79Leu/Phe/Tyr in ParC and Ser81Ile/Phe/Tyr in GyrA were introduced into the R6 background, resulting in 15 isogenic strains. Their level of fluoroquinolone resistance was determined by susceptibility testing for ciprofloxacin, levofloxacin, moxifloxacin, gatifloxacin, gemifloxacin, garenoxacin, and norfloxacin. Leu79 and Ile81 alone as well as 79/81Phe/Tyr substitutions did not contribute significantly to resistance, with fluoroquinolone MICs increasing two- to fourfold compared to wild type for all agents tested. Fluoroquinolone MICs for double transformants ParC Ser79Phe/Tyr/Leu-GyrA Ser81Phe/Tyr were uniformly increased by 8- to 64-fold regardless of pairs of amino acid substitutions. However, combinations including Ile81 conferred two- to fourfold-higher levels of resistance than did combinations including any other Ser81 GyrA substitution, thus demonstrating the differential effects of diverse amino acid substitutions at particular hotspots on fluoroquinolone MICs.

scholarly journals Mutations in the gyrA, parC, and parE Genes Associated with Fluoroquinolone Resistance in Clinical Isolates of Mycoplasma hominis

1999 ◽  
Vol 43 (4) ◽  
pp. 954-956 ◽  
Author(s):  
Cécile M. Bebear ◽  
Joel Renaudin ◽  
Alain Charron ◽  
Hélène Renaudin ◽  
Bertille de Barbeyrac ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mycoplasma Hominis ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Amino Acid Substitutions ◽  
Topoisomerase Iv

ABSTRACT Five clinical isolates of Mycoplasma hominis from three different patients were examined for resistance to fluoroquinolones; some of these isolates were probably identical. All five isolates harbored amino acid substitutions in the quinolone resistance-determining regions of both DNA gyrase (GyrA) and topoisomerase IV (ParC or ParE). Furthermore, the novobiocin MIC for three isolates showed a significant increase. This is the first characterization of fluoroquinolone-resistant clinical mycoplasma isolates from humans.

scholarly journals In Vitro Activities of Novel Nonfluorinated Quinolones PGE 9262932 and PGE 9509924 against Clinical Isolates of Staphylococcus aureus and Streptococcus pneumoniae with Defined Mutations in DNA Gyrase and Topoisomerase IV

2002 ◽  
Vol 46 (6) ◽  
pp. 1651-1657 ◽  
Author(s):  
Mark E. Jones ◽  
Ian A. Critchley ◽  
James A. Karlowsky ◽  
Renée S. Blosser-Middleton ◽  
Franz-Josef Schmitz ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Streptococcus Pneumoniae ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Wild Type ◽  
Topoisomerase Iv ◽  
Selection Of

ABSTRACT Two 8-methoxy nonfluorinated quinolones (NFQs), PGE 9262932 and PGE 9509924, were tested against contemporary clinical isolates of Staphylococcus aureus (n = 122) and Streptococcus pneumoniae (n = 69) with genetically defined quinolone resistance-determining regions (QRDRs). For S. aureus isolates with wild-type (WT) sequences at the QRDRs, the NFQs demonstrated activities 4- to 32-fold more potent (MICs at which 90% of isolates are inhibited [MIC90s], 0.03 μg/ml) than those of moxifloxacin (MIC90, 0.12 μg/ml), gatifloxacin (MIC90, 0.25 μg/ml), levofloxacin (MIC90, 0.25 μg/ml), and ciprofloxacin (MIC90, 1 μg/ml). Against S. pneumoniae isolates with WT sequences at gyrA and parC, the NFQs PGE 9262932 (MIC90, 0.03 μg/ml) and PGE 9509924 (MIC90, 0.12 μg/ml) were 8- to 64-fold and 2- to 16-fold more potent, respectively, than moxifloxacin (MIC90, 0.25 μg/ml), gatifloxacin (MIC90, 0.5 μg/ml), levofloxacin (MIC90, 2 μg/ml), and ciprofloxacin (MIC90, 2 μg/ml). The MICs of all agents were elevated for S. aureus isolates with alterations in GyrA (Glu88Lys or Ser84Leu) and GrlA (Ser80Phe) and S. pneumoniae isolates with alterations in GyrA (Ser81Phe or Ser81Tyr) and ParC (Ser79Phe or Lys137Asn). Fluoroquinolone MICs for S. aureus strains with double alterations in GyrA combined with double alterations in GrlA were ≥32 μg/ml, whereas the MICs of the NFQs for strains with these double alterations were 4 to 8 μg/ml. The PGE 9262932 and PGE 9509924 MICs for the S. pneumoniae isolates did not exceed 0.5 and 1 μg/ml, respectively, even for isolates with GyrA (Ser81Phe) and ParC (Ser79Phe) alterations, for which levofloxacin MICs were >16 μg/ml. No difference in the frequency of selection of mutations (<10−8 at four times the MIC) in wild-type or first-step mutant isolates of S. aureus or S. pneumoniae was detected for the two NFQs. On the basis of their in vitro activities, these NFQ agents show potential for the treatment of infections caused by isolates resistant to currently available fluoroquinolones.

scholarly journals Prevalence, Characteristics, and Molecular Epidemiology of Macrolide and Fluoroquinolone Resistance in Clinical Isolates of Streptococcus pneumoniae at Five Tertiary-Care Hospitals in Korea

2007 ◽  
Vol 51 (7) ◽  
pp. 2625-2627 ◽  
Author(s):  
Jeong Hwan Shin ◽  
Hee Jung Jung ◽  
Hye Ran Kim ◽  
Joseph Jeong ◽  
Seok Hoon Jeong ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Molecular Epidemiology ◽  
Tertiary Care ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
New Mutation ◽  
Tertiary Care Hospitals

ABSTRACT The genes erm(B), mef(A), and both erm(B) and mef(A) were identified in 42.6, 10.1, and 47.3%, respectively, of the erythromycin-resistant Streptococcus pneumoniae isolates. Of the strains, 3.8% were nonsusceptible to levofloxacin and had 1 to 6 amino acid changes in the quinolone resistance-determining region, including a new mutation, Asn94Ser, in the product of parC. Levofloxacin with reserpine was highly specific for efflux screening.

Involvement of topoisomerase IV and DNA gyrase as ciprofloxacin targets in Streptococcus pneumoniae.

1996 ◽  
Vol 40 (10) ◽  
pp. 2321-2326 ◽  
Author(s):  
X S Pan ◽  
J Ambler ◽  
S Mehtar ◽  
L M Fisher
Keyword(s):  
Streptococcus Pneumoniae ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Resistant Isolate ◽  
Topoisomerase Iv ◽  
E Coli ◽  
Low Level ◽  
Drug Concentrations ◽  
Level Resistance

Ciprofloxacin-resistant mutants of Streptococcus pneumoniae 7785 were generated by stepwise selection at increasing drug concentrations. Sequence analysis of PCR products from the strains was used to examine the quinolone resistance-determining regions of the GyrA and GyrB proteins of DNA gyrase and the analogous regions of the ParC and ParE subunits of DNA topoisomerase IV. First-step mutants exhibiting low-level resistance had no detectable changes in their topoisomerase quinolone resistance-determining regions, suggesting altered permeation or another novel resistance mechanism. Nine of 10 second-step mutants exhibited an alteration in ParC at Ser-79 to Tyr or Phe or at Ala-84 to Thr. Third- and fourth-step mutants displaying high-level ciprofloxacin resistance were found to have, in addition to the ParC alteration, a change in GyrA at residues equivalent to Escherichia coli GyrA resistance hot spots Ser-83 and Asp-87 or in GyrB at Asp-435 to Asn, equivalent to E. coli Asp-426, part of a highly conserved EGDSA motif in GyrB. No ParE changes were observed. Complementary analysis of two S. pneumoniae clinical isolates displaying low-level resistance to ciprofloxacin revealed a ParC change at Ser-79 to Phe or Arg-95 to Cys but no changes in GyrA, GyrB, or ParE. A highly resistant isolate, in addition to a ParC mutation, had a GyrA alteration at the residue equivalent to E. coli Asp-87. Thus, in both laboratory strains and clinical isolates, ParC mutations preceded those in GyrA, suggesting that topoisomerase IV is a primary topoisomerase target and gyrase is a secondary target for ciprofloxacin in S. pneumoniae.

scholarly journals Alterations in the GyrA subunit of DNA gyrase and the ParC subunit of topoisomerase IV in quinolone-resistant clinical isolates of Klebsiella pneumoniae.

1997 ◽  
Vol 41 (3) ◽  
pp. 699-701 ◽  
Author(s):  
T Deguchi ◽  
A Fukuoka ◽  
M Yasuda ◽  
M Nakano ◽  
S Ozeki ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Primary Target ◽  
Gyra Gene ◽  
Topoisomerase Iv ◽  
Complementary Role

We determined a partial sequence of the Klebsiella pneumoniae parC gene, including the region analogous to the quinolone resistance-determining region of the Escherichia coli gyrA gene, and examined 26 clinical strains of K. pneumoniae for an association of alterations in GyrA and ParC with susceptibilities to quinolones. The study suggests that in K. pneumoniae DNA gyrase is a primary target of quinolones and that ParC alterations play a complementary role in the development of higher-level fluoroquinolone resistance.

scholarly journals Type II Topoisomerase Mutations in Fluoroquinolone-Resistant Clinical Strains of Pseudomonas aeruginosa Isolated in 1998 and 1999: Role of Target Enzyme in Mechanism of Fluoroquinolone Resistance

2001 ◽  
Vol 45 (8) ◽  
pp. 2263-2268 ◽  
Author(s):  
Takaaki Akasaka ◽  
Mayumi Tanaka ◽  
Akihito Yamaguchi ◽  
Kenichi Sato
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Amino Acid ◽  
Dna Gyrase ◽  
Site Directed Mutagenesis ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Type Ii ◽  
Topoisomerase Iv ◽  
Major Mechanism ◽  
Inhibitory Activities

ABSTRACT The major mechanism of resistance to fluoroquinolones forPseudomonas aeruginosa is the modification of type II topoisomerases (DNA gyrase and topoisomerase IV). We examined the mutations in quinolone-resistance-determining regions (QRDR) ofgyrA, gyrB, parC, and parE genes of recent clinical isolates. There were 150 isolates with reduced susceptibilities to levofloxacin and 127 with reduced susceptibilities to ciprofloxacin among 513 isolates collected during 1998 and 1999 in Japan. Sequencing results predicted replacement of an amino acid in the QRDR of DNA gyrase (GyrA or GyrB) for 124 of the 150 strains (82.7%); among these, 89 isolates possessed mutations in parC orparE which lead to amino acid changes. Substitutions of both Ile for Thr-83 in GyrA and Leu for Ser-87 in ParC were the principal changes, being detected in 48 strains. These replacements were obviously associated with reduced susceptibilities to levofloxacin, ciprofloxacin, and sparfloxacin; however, sitafloxacin showed high activity against isolates with these replacements. We purified GyrA (The-83 to Ile) and ParC (Ser-87 to Leu) by site-directed mutagenesis and compared the inhibitory activities of the fluoroquinolones. Sitafloxacin showed the most potent inhibitory activities against both altered topoisomerases among the fluoroquinolones tested. These results indicated that, compared with other available quinolones, sitafloxacin maintained higher activity against recent clinical isolates with multiple mutations ingyrA and parC, which can be explained by the high inhibitory activities of sitafloxacin against both mutated enzymes.

scholarly journals Alterations in the GyrA Subunit of DNA Gyrase and the ParC Subunit of DNA Topoisomerase IV Associated with Quinolone Resistance in Enterococcus faecalis

1998 ◽  
Vol 42 (2) ◽  
pp. 433-435 ◽  
Author(s):  
Emiko Kanematsu ◽  
Takashi Deguchi ◽  
Mitsuru Yasuda ◽  
Takeshi Kawamura ◽  
Yoshinori Nishino ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Primary Target ◽  
Topoisomerase Iv ◽  
Dna Topoisomerase ◽  
High Level

ABSTRACT The gyrA and parC genes of 31 clinical isolates of Enterococcus faecalis, including fluoroquinolone-resistant isolates, were partially sequenced and analyzed for target alterations. Topoisomerase IV may be a primary target in E. faecalis, but high-level fluoroquinolone resistance was associated with simultaneous alterations in both GyrA and ParC.

scholarly journals Impact of Amino Acid Substitutions in B Subunit of DNA Gyrase in Mycobacterium leprae on Fluoroquinolone Resistance

2012 ◽  
Vol 6 (10) ◽  
pp. e1838 ◽  
Author(s):  
Kazumasa Yokoyama ◽  
Hyun Kim ◽  
Tetsu Mukai ◽  
Masanori Matsuoka ◽  
Chie Nakajima ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mycobacterium Leprae ◽  
Dna Gyrase ◽  
Fluoroquinolone Resistance ◽  
Amino Acid Substitutions ◽  
B Subunit

scholarly journals Relationship between Mutations in the gyrA Gene and Quinolone Resistance in Clinical Isolates of Corynebacterium striatum and Corynebacterium amycolatum

2005 ◽  
Vol 49 (5) ◽  
pp. 1714-1719 ◽  
Author(s):  
Josep M. Sierra ◽  
Luis Martinez-Martinez ◽  
Fernando Vázquez ◽  
Ernest Giralt ◽  
Jordi Vila
Keyword(s):  
Amino Acid ◽  
Test Method ◽  
Quinolone Resistance ◽  
Clinical Isolates ◽  
Fluoroquinolone Resistance ◽  
Moderate Increase ◽  
Gyra Gene ◽  
Double Mutation ◽  
Corynebacterium Striatum

ABSTRACT Quinolone susceptibility was analyzed in 17 clinical isolates of Corynebacterium striatum and 9 strains of Corynebacterium amycolatum by the E-test method in Mueller-Hinton agar plates. The C. striatum ATCC 6940 strain was used as a control strain. The amplified quinolone resistance determining regions of the gyrA genes of C. amycolatum and C. striatum were characterized. Four in vitro quinolone-resistant mutants of C. amycolatum were selected and analyzed. Both in vivo and in vitro quinolone-resistant strains of C. amycolatum showed high levels of fluoroquinolone resistance in strains with a double mutation leading to an amino acid change in positions 87 and 91 or positions 87 and 88 (unusual mutation) of GyrA, whereas the same concomitant mutations at amino acid positions 87 and 91 in GyrA of C. striatum produced high levels of resistance to ciprofloxacin and levofloxacin but only showed a moderate increase in the MIC of moxifloxacin, suggesting that other mechanism(s) of quinolone resistance could be involved in moxifloxacin resistance in C. amycolatum. Moreover, a PCR-RFLP-NcoI of the gyrA gene was developed to distinguish between C. amycolatum and C. striatum species.

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