High Levels Of Quinolone Resistance Research Articles

PREVALENCE OF FLUOROQUINOLONES RESISTANCE AMONG ESBL-PRODUCING ESCHERICHIA COLI ISOLATED FROM URINE SAMPLES

BACKGROUND
 Resistance among uropathogens has emerged as a growing concern necessitating re-evaluation of the efficacy of recommended empiric antimicrobial regimens. Misuse and overuse of various antibacterial agents in hospitals is a key cause of the emergence of anti-microbial resistance.
 Aims: To identify the prevalence of extended-spectrum beta-lactamase (ESBL) producing Escherichia coli from urine specimens and to know the prevalence of Fluoroquinolone resistance among ESBL-producing Escherichia coli (E. coli) isolates.
 Material and method: The study was conducted on 500 E. coli isolates from urine samples received in the Department of Microbiology, MMIMSR, Mullana, Ambala. The organism isolation and identification were done as per the standard procedures and antibiotic sensitivity testing was done following CLSI guidelines. All the strains were screened out for ESBL production as per CLSI guidelines. Ciprofloxacin, Norfloxacin, and Ofloxacin discs were used for the detection of fluoroquinolone resistance.
 Results: Out of the total 500 E. Coli isolates from urine samples received in our laboratory, 69% were found to be ESBL producers.
 Conclusion: We found a high level of quinolone resistance among ESBL-producing Escherichia coli strains isolated from patients with UTI. Because of the widespread use of fluoroquinolones in our country, there is a need for sensitive antibiotic stewardship. Further research is needed to ascertain the gravity of quinolone resistance and to swiftly act against its spread among other nosocomial pathogens.

High frequency of mutations in gyrA gene associated with quinolones resistance in uropathogenic Escherichia coli isolates from the north of Iran.

Objective(s):Regarding the global burden of uropathogenic Escherichia coli (UPEC) infections, prevention and treatment of such infections play a significant role in healthcare management. The inordinate use of fluoroquinolones led to a worldwide spread of quinolone-resistant strains. Therefore, this study aimed to investigate mutations in codons 83 and 106 of gyrA gene in UPEC isolates in the north of Iran.Materials and Methods:This cross-sectional study performed on a total of 223 UPEC isolates which were recovered within 6 months in 2017. Isolates were identified and confirmed by standard microbiologic tests, and antimicrobial susceptibility testing was carried out by disk diffusion and E-test methods. PCR reaction was performed to amplify gyrA gene, and PCR-RFLP was performed using BsiEI and BstUI restriction enzymes to investigate mutations in gyrA gene.Results:The nalidixic acid, ciprofloxacin, ofloxacin, and norfloxacin resistance rates were 61.9%, 50.2%, 48.25, and 45.3%, respectively. Overall, 55.2% of E. coli isolates had a mutation in gyrA gene in codon 83, and 20.2% in codon 106. Also, 15.2% of isolates had simultaneously mutation. Moreover, a significant association was found between mutations in gyrA gene and quinolone and fluoroquinolones resistance pattern of UPEC isolates.Conclusion:Our results revealed a high level of quinolone resistance associated with the mutations in gyrA among the clinical isolates of UPEC in our region. To the best of our knowledge, this study is the first investigation on the role of gyrA alteration in quinolone resistance among UPEC isolates from the north of Iran.

Quinolone Resistance Mechanisms Among Salmonella enterica in Malaysia.

The prevalence of quinolone-resistant Salmonella enterica is on the rise worldwide. Salmonella enterica is one of the major foodborne pathogens in Malaysia. Therefore, we aim to investigate the occurrence and mechanisms of quinolone resistance among Salmonella strains isolated in Malaysia. A total of 283 Salmonella strains isolated from food, humans, and animals were studied. The disk diffusion method was used to examine the quinolone susceptibility of the strains, and the minimum inhibitory concentration (MIC) values of nalidixic acid and ciprofloxacin were also determined. DNA sequencing of the quinolone resistance-determining regions (QRDRs) of gyrase and topoisomerase IV genes and the plasmid-borne qnr genes was performed. The transfer of the qnr gene was examined through transconjugation experiment. A total of 101 nalidixic acid-resistant Salmonella strains were identified. In general, all strains were highly resistant to nalidixic acid (average MICNAL, 170 μg/ml). Resistance to ciprofloxacin was observed in 30.7% of the strains (1 ≤ MICCIP ≤ 2 μg/ml). Majority of the strains contained missense mutations in the QRDR of gyrA (69.3%). Silent mutations were frequently detected in gyrB (75.2%), parC (27.7%), and parE (51.5%) within and beyond the QRDRs. Novel mutations were detected in parC and parE. The plasmid-borne qnrS1 variant was found in 36.6% of the strains, and two strains were found to be able to transfer the qnrS1 gene. Overall, mutations in gyrA and the presence of qnrS1 genes might have contributed to the high level of quinolone resistance among the strains. Our study provided a better understanding on the status of quinolone resistance among Salmonella strains circulating in Malaysia.

Niveles de resistencia a quinolonas y otros antimicrobianos en cepas de Escherichia coli comensales en niños de la zona periurbana de Lima, Perú

The main aim of this study was to establish the resistance levels to antimicrobial agents, in 222 non-pathogenic E. Coli strains of fecal origin in Peru. The proportion of resistance found to the evaluated antimicrobials was ampicillin (62.6%), cotrimoxazole (48,6%), tetracycline (43,0%) and chloramphenicol (15,8%). We emphasize the high resistance levels found for quinolones: 32% for nalidixic acid (NAL) and 12% for ciprofloxacin (CIP). These high levels of quinoloneresistance in non-pathogenic strains isolated from children in this age group highlight the extensive use and the impact of the intake of this kind of antimicrobials in the community, showing the potential risk of the loss of their utility in the area.

Molecular characterization of fluoroquinolone-resistant Aeromonas spp. isolated from imported shrimp.

Sixty-three nalidixic acid-resistant Aeromonas sp. isolates were obtained from imported shrimp. Phylogenetic analysis of gyrB sequences indicated that 18 were A. enteropelogenes, 26 were A. caviae, and 19 were A. sobria. Double missense mutations in the quinolone resistance-determining region (QRDR) of gyrA at codon 83 (Ser→Val/Ile) and codon 92 (Leu→Met) coupled with a point mutation of parC at codon 80 (Ser→Ile/Phe) conferred high levels of quinolone resistance in the isolates. A majority of A. enteropelogenes and A. caviae strains harbored toxin genes, whereas only a few A. sobria strains harbored these genes. The fluoroquinolone-resistant Aeromonas spp. exhibited higher cytotoxicity than fluoroquinolone-sensitive, virulent Aeromonas spp. to rat epithelial cells.

Niveles de resistencia a quinolonas y otros antimicrobianos en cepas de Escherichia coli comensales en niños de la zona periurbana de Lima, Perú

The main aim of this study was to establish the resistance levels to antimicrobial agents, in 222 non-pathogenic E. coli strains of fecal origin in Peru. The proportion of resistance found to the evaluated antimicrobials was ampicillin (62.6%), cotrimoxazole (48,6%), tetracycline (43,0%) and chloramphenicol (15,8%). We emphasize the high resistance levels found for quinolones: 32% for nalidixic acid (NAL) and 12% for ciprofloxacin (CIP). These high levels of quinoloneresistance in non-pathogenic strains isolated from children in this age group highlight the extensive use and the impact of the intake of this kind of antimicrobials in the community, showing the potential risk of the loss of their utility in the area.

Emergence of Proteus mirabilis Harboring bla KPC-2 and qnrD in a Chinese Hospital

Nineteen carbapenem-nonsusceptible Proteus mirabilis isolates were recovered from intensive care units in the Second Affiliated Hospital of Zhejiang University during a 3-month period. The isolates showed a high level of resistance against ciprofloxacin, in addition to their resistance against the carbapenems. Pulsed-field gel electrophoresis (PFGE) analysis showed that these isolates belonged to three clonal strains. PCRs and DNA sequence analysis of the carbapenemase and other β-lactamase genes indicated that all the isolates harbored the bla(KPC-2) gene. Twelve of 19 isolates harbored the plasmid-mediated quinolone resistance (PMQR) genes, both the qnrD and aac(6')-Ib-cr genes. Eight representative isolates with high levels of quinolone resistance carried the similar mutation profiles of S83I in gyrA, E466D in gyrB, and S80I in parC. Reduced carbapenem susceptibility was transferred to Escherichia coli (EC600) in a conjugation experiment, while the quinolone resistance was not. DNA hybridization showed that qnrD was located on a plasmid of approximately 4.5 kb. In summary, large clonally related isolates of KPC-2-producing P. mirabilis emerged in a Chinese hospital, and qnrD was detected in KPC-producing P. mirabilis for the first time.

High level of quinolone resistance in Escherichia coli from healthy chicken broilers

The development of resistance to quinolones (nalidixic acid, ciprofloxacin and enrofloxacin) in 2006-2008 was evaluated in 317 strains of Escherichia coli isolated from healthy chicken broilers from various farms. The isolates (2006/2007/2008) showed a high resistance to nalidixic acid (87/85/67 %), ciprofloxacin (CIP) (49/54/29 %) and enrofloxacin (ENR) (52/42/22 %). Nalidixic acid-resistant isolates with low level of MIC for CIP and ENR represented a single mutation; intermediary MIC for CIP and ENR were related to two mutations and high level resistance MIC for CIP (> or =4 mg/L) and ENR (> or =16 mg/L) represented three mutations (two in gyrA and one in parC). There was a correlation between the phenotype reading of high-level resistance and mutations in gyrA (Ser83Leu, Asp87Tyr or Asp87Asn) and parC (Ser80Ile) gene. Plasmid-mediated quinolone-resistance qnrS gene was detected in one Escherichia coli strain with a high level of ciprofloxacin resistance. Our results demonstrate the increase in occurrence of multiresistant E. coli strains with a high level of chromosomal and plasmid resistance to fluoroquinolones.

Mutant Prevention Concentrations of Fluoroquinolones for Enterobacteriaceae Expressing the Plasmid-Carried Quinolone Resistance Determinant qnrA1

The influence of qnrA1 on the development of quinolone resistance in Enterobacteriaceae was evaluated by using the mutant prevention concentration parameter. The expression of qnrA1 considerably increased the mutant prevention concentration compared to strains without this gene. In the presence of qnrA1, mutations in gyrA and parC genes were easily selected to produce high levels of quinolone resistance.

Clonal Expansion May Account for High Levels of Quinolone Resistance in Salmonella enterica Serovar Enteritidis

We have observed a high incidence of isolated nalidixic acid resistance in Salmonella enterica serovar Enteritidis isolates in Ireland, particularly isolates of phage type 1 (PT1). A group of nalidixic acid-resistant (n = 22) and nalidixic acid-susceptible (n = 28) isolates of serovar Enteritidis from multiple sites in Ireland were selected. Isolates were typed by pulsed-field gel electrophoresis (PFGE) with XbaI, and the MICs for nalidixic acid and ciprofloxacin were determined. Mutations associated with nalidixic acid resistance in clinical isolates and laboratory mutants of serovar Enteritidis and 32 nalidixic acid-resistant isolates of 15 other salmonella serovars were identified. PFGE had limited discriminatory power. A specific point mutation (G246T) associated with amino acid substitution Asp87Tyr in the quinolone resistance determining region of the gyrA gene accounted for 95% of all mutations in serovar Enteritidis and for all mutations in PT1 isolates. Greater diversity of mutations was observed among all non-Enteritidis salmonella serovars studied. Rates of nalidixic acid resistance in serovar Enteritidis may predominantly reflect clonal expansion after infrequent mutation or selection events.

An unusually high level of quinolone resistance associated with type II topoisomerase mutations in quinolone resistance-determining regions of Aeromonas caviae isolated from diarrhoeal patients

We examined 158 strains belonging to different Aeromonas species isolated from hospitalized acute diarrhoea cases for susceptibility to quinolones. Compared to other species, a high percentage of the A. caviae strains were resistant to ciprofloxacin and norfloxacin. Based on MIC values, 6 A. caviae strains were selected and the nucleotide sequences for the quinolone-resistant-determining regions (QRDRs) of gyrA, gyrB and parC genes were analysed. In resistant strains, double mutations (Ser 83→Ile and Asp 87→Asn) and a single mutation (Ser 80→Ile) were detected in the QRDR of gyrA and parC, respectively.

A Mutation in Yeast TOP2 Homologous to a Quinolone-resistant Mutation in Bacteria: MUTATION OF THE AMINO ACID HOMOLOGOUS TO Ser83 OF ESCHERICHIA COLI gyrA ALTERS SENSITIVITY TO EUKARYOTIC TOPOISOMERASE INHIBITORS

In prokaryotic type II topoisomerases (DNA gyrases), mutations that result in resistance to quinolones frequently occur at Ser83 or Ser84 of the gyrA subunit. Mutations to Trp, Ala, and Leu have been identified, all of which confer high levels of quinolone resistance. Extensive segments of DNA gyrase are homologous to eukaryotic topoisomerase II, and Ser741 of yeast TOP2 is homologous to Ser83 of prokaryotic DNA gyrA. Introduction of the Ser741-->Trp mutation into yeast TOP2 confers resistance to 6,8-difluoro-7-(4'-hydroxyphenyl)-1-cyclopropyl- 4-quinolone-3-carboxylic acid (CP-115,953), a fluoroquinolone with substantial activity against eukaryotic topoisomerase II, whereas changing Ser741 to either Leu or Ala does not change sensitivity to quinolones. Interestingly, Ser741-->Trp in the yeast TOP2 also confers hypersensitivity to etoposide. Sensitivity to intercalating anti-topoisomerase II agents such as amsacrine is not changed by any of the three mutations. The topoisomerase II protein carrying the Ser741-->Trp mutation was overexpressed and purified. The purified mutant enzyme had enhanced levels of etoposide stabilized covalent complex as compared with the wild type enzyme and reduced cleavage with CP-115,953. Unlike the wild type enzyme, etoposide-stabilized cleavage is not readily reversible by heat. We suggest that Ser741 is near a binding site for both quinolones and etoposide and that the Ser741-->Trp mutation leads to a more stable ternary complex between etoposide, DNA, and the mutant enzyme.

Quinolone action in Escherichia coli cells carrying gyrA and gyrB mutations.

We have isolated spontaneous mutant strains of Escherichia coli KL16 showing different levels of nalidixic acid (NAL) resistance. From 40 independent mutants, 36 had gyrA and four had gyrB mutations. Most of the gyrA mutations (30/36) conferred high level NAL resistance. In contrast, the only gyrB mutation that conferred a relatively high level of NAL resistance also determined enhanced susceptibility to quinolones with a piperazinyl substituent at C7 position of the quinolone ring (amphoteric quinolones). This gyrB mutation (denoted gyrB1604), jointly with a gyrA mutation (denoted gyrA972) which confers a high level of quinolone resistance, were used to construct strain IC2476, carrying the two gyr mutant alleles. The susceptibility of this strain to amphoteric quinolones (pipemidic acid, norfloxacin and ciprofloxacin) was similar to that of the gyrA972 single mutant. This result indicates that the change in GyrA subunit which determines a high level of quinolone-resistance has the capacity to mask the hypersusceptibility to amphoteric quinolones promoted by the GyrB1604 mutant subunit. This capacity was further confirmed by studying the effects of ciprofloxacin (CFX) on gyrase inhibition in the gyrA972 gyrB1604 strain.

Quinolone action inEscherichia colicells carryinggyrAandgyrBmutations

We have isolated spontaneous mutant strains of Escherichia coli KL16 showing different levels of nalidixic acid (NAL) resistance. From 40 independent mutants, 36 had gyrA and four had gyrB mutations. Most of the gyrA mutations (30/36) conferred high level NAL resistance. In contrast, the only gyrB mutation that conferred a relatively high level of NAL resistance also determined enhanced susceptibility to quinolones with a piperazinyl substituent at C7 position of the quinolone ring (amphoteric quinolones). This gyrB mutation (denoted gyrB1604), jointly with a gyrA mutation (denoted gyrA972) which confers a high level of quinolone resistance, were used to construct strain IC2476, carrying the two gyr mutant alleles. The susceptibility of this strain to amphoteric quinolones (pipemidic acid, norfloxacin and ciprofloxacin) was similar to that of the gyrA972 single mutant. This result indicates that the change in GyrA subunit which determines a high level of quinolone-resistance has the capacity to mask the hypersusceptibility to amphoteric quinolones promoted by the GyrB1604 mutant subunit. This capacity was further confirmed by studying the effects of ciprofloxacin (CFX) on gyrase inhibition in the gyrA972 gyrB1604 strain.

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 OmpF reduction.

Chromosomal multiple-antibiotic-resistant (Mar) mutants of Escherichia coli, selected on agar containing low concentrations of tetracycline or chloramphenicol, were 6- to 18-fold less susceptible to the fluoroquinolones than were their wild-type E. coli K-12 or E. coli C parental strains. The frequency of emergence of such mutants was at least 1,000-fold higher than that of those selected by the fluoroquinolone norfloxacin directly. When Mar mutants, but not wild-type cells, were plated on norfloxacin, mutants resistant to high levels of norfloxacin (2 micrograms/ml) appeared at a relatively high (approximately 10(-7] frequency. In addition to decreased amounts of OmpF, Mar mutants had other outer membrane protein changes and were four- to eightfold less susceptible to fluoroquinolones than was an ompF::Tn5 mutant lacking only OmpF. Accumulation of [3H]norfloxacin was more than threefold lower in the Mar mutants than in wild-type cells and twofold lower than in the OmpF-deficient derivative. These differences were not attributable to a change in the endogenous active efflux system for norfloxacin in E. coli. Norfloxacin-induced inhibition of DNA synthesis was threefold lower in intact cells of a Mar mutant than in susceptible cells, but this difference was not seen in toluene-permeabilized cells. Insertion of Tn5 into marA (min 34.05 on the chromosome) led to a return of the wild-type patterns of norfloxacin accumulation, fluoroquinolone and other antimicrobial agent susceptibilities, and outer membrane protein profile, including partial restoration of OmpF. These findings together suggest that marA-dependent fluoroquinolone resistance is linked to decreased cell permeability, only part of which can be accounted for by the reduction in OmpF. Once mutated to marA, cells can achieve high levels of quinolone resistance at a relatively high frequency.

Methicillin-resistant strains of Staphylococcus aureus resistant to quinolones

Since January 1988, Staphylococcus aureus strains with a high level of quinolone resistance have been isolated at 17 hospitals and 15 nursing homes in New York City. The majority of these strains were methicillin resistant. The bacteriophage types and susceptibility to other antibiotics were similar to those of quinolone-susceptible strains isolated at the same hospitals.