Quinolone Resistance Mechanisms Research Articles

A Review of the Impact of Streptococcal Infections and Antimicrobial Resistance on Human Health.

Streptococcus pneumoniae, Streptococcus pyogenes (GAS), and Streptococcus agalactiae (GBS) are bacteria that can cause a range of infections, some of them life-threatening. This review examines the spread of antibiotic resistance and its mechanisms against antibiotics for streptococcal infections. Data on high-level penicillin-resistant invasive pneumococci have been found in Brazil (42.8%) and Japan (77%). The resistance is caused by mutations in genes that encode penicillin-binding proteins. Similarly, GAS and GBS strains reported from Asia, the USA, and Africa have undergone similar transformations in PBPs. Resistance to major alternatives of penicillins, macrolides, and lincosamides has become widespread among pneumococci and streptococci, especially in Asia (70-95%). The combination of several emm types with erm(B) is associated with the development of high-level macrolide resistance in GAS. Major mechanisms are ribosomal target modifications encoded by erm genes, ribosomal alterations, and active efflux pumps that regulate antibiotic entry due to mefA/E and msrD genes. Tetracycline resistance for streptococci in different countries varied from 22.4% in the USA to 83.7/100% in China, due to tet genes. Combined tetracycline/macrolide resistance is usually linked with the insertion of ermB into the transposon carrying tetM. New quinolone resistance is increasing by between 11.5 and 47.9% in Asia and Europe. The mechanism of quinolone resistance is based on mutations in gyrA/B, determinants for DNA gyrase, or parC/E encoding topoisomerase IV. The results for antibiotic resistance are alarming, and urgently call for increased monitoring of this problem and precautionary measures for control to prevent the spread of resistant mutant strains.

Impact of suppression of the SOS response on protein expression in clinical isolates of Escherichia coli under antimicrobial pressure of ciprofloxacin.

Suppression of the SOS response in combination with drugs damaging DNA has been proposed as a potential target to tackle antimicrobial resistance. The SOS response is the pathway used to repair bacterial DNA damage induced by antimicrobials such as quinolones. The extent of lexA-regulated protein expression and other associated systems under pressure of agents that damage bacterial DNA in clinical isolates remains unclear. The aim of this study was to assess the impact of this strategy consisting on suppression of the SOS response in combination with quinolones on the proteome profile of Escherichia coli clinical strains. Five clinical isolates of E. coli carrying different chromosomally- and/or plasmid-mediated quinolone resistance mechanisms with different phenotypes were selected, with E. coli ATCC 25922 as control strain. In addition, from each clinical isolate and control, a second strain was created, in which the SOS response was suppressed by deletion of the recA gene. Bacterial inocula from all 12 strains were then exposed to 1xMIC ciprofloxacin treatment (relative to the wild-type phenotype for each isogenic pair) for 1 h. Cell pellets were collected, and proteins were digested into peptides using trypsin. Protein identification and label-free quantification were done by liquid chromatography-mass spectrometry (LC-MS) in order to identify proteins that were differentially expressed upon deletion of recA in each strain. Data analysis and statistical analysis were performed using the MaxQuant and Perseus software. The proteins with the lowest expression levels were: RecA (as control), AphA, CysP, DinG, DinI, GarL, PriS, PsuG, PsuK, RpsQ, UgpB and YebG; those with the highest expression levels were: Hpf, IbpB, TufB and RpmH. Most of these expression alterations were strain-dependent and involved DNA repair processes and nucleotide, protein and carbohydrate metabolism, and transport. In isolates with suppressed SOS response, the number of underexpressed proteins was higher than overexpressed proteins. High genomic and proteomic variability was observed among clinical isolates and was not associated with a specific resistant phenotype. This study provides an interesting approach to identify new potential targets to combat antimicrobial resistance.

Molecular mechanisms of quinolone resistance in non-typhoidal Salmonella

By conducting retrospective analysis, this study aim to investigate the resistance mechanism of quinolones in non-typhoidal Salmonella (NTS). A total of 105 strains of NTS isolated from clinical specimens from the Fifth Affiliated Hospital of Southern Medical University from May 2020 to February 2021 were used as research objects. VITEK2 Compact automatic identification drug sensitivity analysis system and serological test were used to identify the strains. The sensitivity of the strains to ciprofloxacin, levofloxacin and nalidixic acid was detected by AGAR dilution method. The whole genome of 105 strains of NTS was sequenced. Abricate and other softwares were used to analyze drug-resistant genes, including plasmid-mediated quinolone resistance gene (PMQR) and Quinolone resistance determination region (QRDR). Serotypes and ST types were analyzed using SISTR and MLST, and phylogenetic trees were constructed. The results showed that the NTS isolated in this region were mainly ST34 Salmonella typhimurium (53.3%). The drug sensitivity results showed that the drug resistance rates of NTS to ciprofloxacin, levofloxacin and nalidixic acid were 30.4%, 1.9% and 22.0%, respectively, and the intermediate rates of ciprofloxacin and levofloxacin were 27.6% and 54.2%.A total of 46 (74.2%) of the 62 quinolone non-susceptible strains carried the PMQR gene, mainly qnrS1 (80.4%), followed by aac(6')-Ib-cr(15.2%); there were 14 NTS and 8 NTS had gyrA and parC gene mutations, respectively. The gyrA was mutations at the amino acid position 87, Asp87Tyr, Asp87Asn, Asp87Gly, and Thr57Ser mutations were detected in parC. In conclusion, this study found that NTS had relatively high resistance to quinolones, carrying qnrS1 gene mainly resulted in decreased sensitivity of NTS to ciprofloxacin and levofloxacin, and gyrA:87 mutation mainly resulted in NTS resistance to Nalidixic acid; Salmonella typhimurium in clinical isolates showed clonal transmission and required further epidemiological surveillance.

Evaluation and Characterization of Quinolone-Resistant Escherichia coli in Wastewater Treatment Plant Effluents

The increasing global incidence of quinolone antimicrobial resistance poses a considerable public health concern. The aquatic environment, particularly wastewater treatment plants (WWTPs), serves as a major reservoir for antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs), leading to the dissemination of antibiotic resistance. This study aimed to assess the prevalence and factors contributing to quinolone antibiotic resistance in Escherichia coli isolates obtained from effluents of 33 WWTPs. A total of 1082 E. coli isolates were analyzed, 32.6% and 17.1% of which showed resistance to nalidixic acid and ciprofloxacin, respectively. Phenotypic and genotypic analyses of antibiotic resistance demonstrated that quinolone resistance primarily originated from chromosomal mutations in the gyrA, parC, and parE genes, known as quinolone resistance-determining regions (QRDRs). The amino acid substitution at codon 83 in gyrA was closely associated with nalidixic acid resistance, whereas substitutions at codon 87 in gyrA and codon 80 in parC were significantly associated with ciprofloxacin resistance. The plasmid-mediated quinolone resistance (PMQR) genes qnrS and qnrB were identified in 41 isolates (11.5%) and 15 isolates (4.2%), respectively. Thus, we confirmed that the quinolone resistance in E. coli in WWTPs primarily occurs through QRDR mutations rather than through the acquisition of PMQR genes. Phylogenetic analysis revealed that most quinolone-resistant isolates belonged to the B1, A, B2, and D phylogenetic groups. Notably, the B2 group, which is responsible for extraintestinal infections, exhibited the highest rate of quinolone resistance. These findings provide novel insights into the presence and mechanisms of quinolone resistance in E. coli isolates from WWTPs, emphasizing the need for further research and understanding of quinolone resistance in the environment.

Systematic investigation of the emerging pathogen of Tsukamurella species in a Chinese tertiary teaching hospital.

Tsukamurella species have been clinically regarded as rare but emerging opportunistic pathogens causing various infections in humans. Tsukamurella pneumonia has often been misdiagnosed as pulmonary tuberculosis due to its clinical presentation resembling tuberculosis-like syndromes. Tsukamurella species have also been confused in the laboratory with other phylogenetic bacteria, such as Gordonia. This study aimed to investigate the clinical, microbiological, and molecular characteristics; species distribution; and antimicrobial susceptibility of Tsukamurella species. Immunodeficiency and chronic pulmonary disease appeared to be risk factors for Tsukamurella pneumonia, and the presence of bronchiectasis and pulmonary nodules on imaging was highly correlated with this infection. The study confirmed that groEL (heat shock protein 60) and secA (the secretion ATPase) genes are reliable for identifying Tsukamurella species. Additionally, the ssrA (stable small RNA) gene showed promise as a tool for discriminating between different Tsukamurella species with the shortest sequence length. In terms of antimicrobial susceptibility, quinolones, trimethoprim/sulfamethoxazole, amikacin, minocycline, linezolid, and tigecycline demonstrated potent in vitro activity against Tsukamurella isolates in our study. The study also proposed a resistance mechanism involving a substitution (S91R) within the quinolone-resistance-determining region of the gyrA gene, which confers resistance to levofloxacin and ciprofloxacin. Furthermore, we found that disk diffusion testing is not suitable for testing the susceptibilities of Tsukamurella isolates to ciprofloxacin, imipenem, and minocycline. In conclusion, our systematic investigation may contribute to a better understanding of this rare pathogen. IMPORTANCE Tsukamurella species are rare but emerging human pathogens that share remarkable similarities with other mycolic acid-containing genera of the order Actinomycetales, especially Mycobacterium tuberculosis. Consequently, misdiagnosis and therapeutic failures can occur in clinical settings. Despite the significance of accurate identification, antimicrobial susceptibility, and understanding the resistance mechanism of this important genus, our knowledge in these areas remains fragmentary and incomplete. In this study, we aimed to address these gaps by investigating promising identification methods, the antimicrobial susceptibility patterns, and a novel quinolone resistance mechanism in Tsukamurella species, utilizing a collection of clinical isolates. The findings of our study will contribute to improve diagnosis and successful management of infections caused by Tsukamurella species, as well as establishing well-defined performance and interpretive criteria for antimicrobial susceptibility testing.

Antimicrobial Susceptibility to 27 Drugs and the Molecular Mechanisms of Macrolide, Tetracycline, and Quinolone Resistance in Gemella sp.

Gemella is a catalase-negative, facultative anaerobic, Gram-positive coccus that is commensal in humans but can become opportunistic and cause severe infectious diseases, such as infective endocarditis. Few studies have tested the antimicrobial susceptibility of Gemella. We tested its antimicrobial susceptibility to 27 drugs and defined the resistant genes using PCR in 58 Gemella strains, including 52 clinical isolates and six type strains. The type strains and clinical isolates included 22 G. morbillorum, 18 G. haemolysans (GH) group (genetically indistinguishable from G. haemolysans and G. parahaemolysans), 13 G. taiwanensis, three G. sanguinis, and two G. bergeri. No strain was resistant to beta-lactams and vancomycin. In total, 6/22 (27.3%) G. morbillorum strains were erythromycin- and clindamycin-resistant ermB-positive, whereas 4/18 (22.2%) in the GH group, 7/13 (53.8%) G. taiwanensis, and 1/3 (33.3%) of the G. sanguinis strains were erythromycin-non-susceptible mefE- or mefA-positive and clindamycin-susceptible. The MIC90 of minocycline and the ratios of tetM-positive strains varied across the different species-G. morbillorum: 2 µg/mL and 27.3% (6/22); GH group: 8 µg/mL and 27.8% (5/18); G. taiwanensis: 8 µg/mL and 46.2% (6/13), respectively. Levofloxacin resistance was significantly higher in G. taiwanensis (9/13 69.2%) than in G. morbillorum (2/22 9.1%). Levofloxacin resistance was associated with a substitution at serine 83 for leucine, phenylalanine, or tyrosine in GyrA. The mechanisms of resistance to erythromycin and clindamycin differed across Gemella species. In addition, the rate of susceptibility to levofloxacin differed across Gemella sp., and the quinolone resistance mechanism was caused by mutations in GyrA alone.

Characterization of Transferrable Mechanisms of Quinolone Resistance (TMQR) among Quinolone-resistant Escherichia coli and Klebsiella pneumoniae causing Urinary Tract Infection in Nepalese Children

BackgroundTransferrable mechanisms of quinolone resistance (TMQR) can lead to fluoroquinolone non-susceptibility in addition to chromosomal mechanisms. Some evidence suggests that fluoroquinolone resistance is increasing among the pediatric population. We sought to determine the occurrence of TMQR genes among quinolone-resistant E. coli and K. pneumoniae causing urinary tract infections among Nepalese outpatient children (< 18 years) and identify molecular characteristics of TMQR-harboring isolates.MethodsWe performed antimicrobial susceptibility testing, phenotypic extended-spectrum β-lactamase (ESBL) and modified carbapenem inactivation method tests, and investigated the presence of six TMQR genes (qnrA, qnrB, qnrS, aac(6’)-Ib-cr, oqxAB, qepA), three ESBL genes (blaCTX−M, blaTEM, blaSHV), and five carbapenemase genes (blaNDM, blaOXA−48, blaKPC, blaIMP, blaVIM). The quinolone resistance-determining region (QRDR) of gyrA and parC were sequenced for 35 TMQR-positive isolates.ResultsA total of 74/147 (50.3%) isolates were TMQR positive by multiplex PCR [aac(6’)-Ib-cr in 48 (32.7%), qnrB in 23 (15.7%), qnrS in 18 (12.3%), qnrA in 1 (0.7%), and oqxAB in 1 (0.7%) isolate]. The median ciprofloxacin minimum inhibitory concentration of TMQR-positive isolates (64 µg/mL) was two-fold higher than those without TMQR (32 µg/mL) (p = 0.004). Ser-83→Leu and Asp-87→Asn in GyrA and Ser-80→Ile in ParC were the most common QRDR mutations (23 of 35). In addition, there was a statistically significant association between TMQR and two β-lactamase genes; blaCTX−M (p = 0.037) and blaTEM (p = 0.000).ConclusionThis study suggests a high prevalence of TMQR among the quinolone-resistant E. coli and K. pneumoniae isolates causing urinary tract infection in children in this area of Nepal and an association with the carriage of ESBL gene. This is a challenge for the management of urinary infections in children. Comprehensive prospective surveillance of antimicrobial resistance in these common pathogens will be necessary to devise strategies to mitigate the emergence of further resistance.

Multidrug Resistance Profiles and Resistance Mechanisms to β-Lactams and Fluoroquinolones in Bacterial Isolates from Hospital Wastewater in Bangladesh.

Multidrug resistance (MDR) is one of the deadliest public health concerns of the 21st century, rendering many powerful antibiotics ineffective. The current study provides important insights into the prevalence and mechanisms of antibiotic resistance in hospital wastewater isolates. In this study, we determined the MDR profile of 68 bacterial isolates collected from five different hospitals in Dhaka, Bangladesh. Of them, 48 bacterial isolates were identified as Enterobacteriaceae. Additionally, we investigated the prevalence and distribution of five beta-lactam resistance genes, as well as quinolone resistance mechanisms among the isolates. The results of this study showed that 87% of the wastewater isolates were resistant to at least three different antibiotic classes, as revealed using the disc diffusion method. Resistance to β-lactams was the most common, with 88.24% of the isolates being resistant, closely followed by macrolides (80.88% resistant). Polymyxin was found to be the most effective against wastewater isolates, with 29.41% resistant isolates. The most common β-lactam resistance genes found in wastewater isolates were blaTEM (76.09%), blaCTX-M1 (71.74%), and blaNDM (67.39%). Two missense mutations in the quinolone resistance-determining region (QRDR) of gyrA (S83L and D87N) and one in both parC (S80I) and parE (S458A) were identified in all isolates, and one in parE (I529L), which had not previously been identified in Bangladesh. These findings suggest that hospital wastewater acts as an important reservoir of antibiotic-resistant bacteria wherein resistance mechanisms to β-lactams and fluoroquinolones are obvious. Our data also emphasize the need for establishing a nationwide surveillance system for antibiotic resistance monitoring to ensure that hospitals sanitize their wastewater before disposal, and regulation to ensure hospital wastewater is kept away from community settings.

Transferable mechanisms of quinolone resistance are more frequent among enterotoxigenic Escherichia coli isolates displaying low-level quinolone resistance.

This study analysed the mechanisms of quinolone resistance among enterotoxigenic Escherichia coli (ETEC) in a periurban area of Lima, Peru. The susceptibility to nalidixic acid and ciprofloxacin, the role of Phe-Arg-b-Naphtylamyde inhibitable-(PAbN) efflux pumps, the presence of mutations in gyrA and parC as well as the presence of aac(6')Ib-cr, qepA, qnrA, qnrB, qnrC, qnrD, qnrVC and oqxAB were determined in 31 ETEC from previous case/control studies of children's diarrhoea. Discordances between disk diffusion, with all isolates showing intermediate or fully resistance to nalidixic acid, and minimal inhibitory concentration (MIC), with 7 isolates being below considered resistance breakpoint, were observed. Twenty-one isolates possessed gyrA mutations (19 S83L, 2 S83A). AAC(6') Ib-cr, QnrS, QnrB and QepA were found in 7, 6, 2 and 1 isolates respectively, with 3 isolates presenting 2 transferable mechanisms of quinolone resistance (TMQR) concomitantly. TMQR were more frequent among isolates with MIC to nalidixic acid ranging from 2 to 16 mg/L (p=0.03), while gyrA mutations were more frequent among isolates with nalidixic acid MIC >= 128 mg/L (p=0.0002). In summary, the mechanisms of quinolone resistance present in ETEC isolates in Peru have been described. Differences in the prevalence of underlying mechanisms associated with final MIC levels were observed. The results suggest two different evolutive strategies to survive in the presence of quinolones related to specific bacterial genetic background.

QUIRMIA-A Phenotype-Based Algorithm for the Inference of Quinolone Resistance Mechanisms in Escherichia coli.

Quinolone resistance in Escherichia coli occurs mainly as a result of mutations in the quinolone-resistance-determining regions of gyrA and parC, which encode the drugs' primary targets. Mutational alterations affecting drug permeability or efflux as well as plasmid-based resistance mechanisms can also contribute to resistance, albeit to a lesser extent. Simplifying and generalizing complex evolutionary trajectories, low-level resistance towards fluoroquinolones arises from a single mutation in gyrA, while clinical high-level resistance is associated with two mutations in gyrA plus one mutation in parC. Both low- and high-level resistance can be detected phenotypically using nalidixic acid and fluoroquinolones such as ciprofloxacin, respectively. The aim of this study was to develop a decision tree based on disc diffusion data and to define epidemiological cut-offs to infer resistance mechanisms and to predict clinical resistance in E. coli. This diagnostic algorithm should provide a coherent genotype/phenotype classification, which separates the wildtype from any non-wildtype and further differentiates within the non-wildtype. Phenotypic susceptibility of 553 clinical E. coli isolates towards nalidixic acid, ciprofloxacin, norfloxacin and levofloxacin was determined by disc diffusion, and the genomes were sequenced. Based on epidemiological cut-offs, we developed a QUInolone Resistance Mechanisms Inference Algorithm (QUIRMIA) to infer the underlying resistance mechanisms responsible for the corresponding phenotypes, resulting in the categorization as "susceptible" (wildtype), "low-level resistance" (non-wildtype) and "high-level resistance" (non-wildtype). The congruence of phenotypes and whole genome sequencing (WGS)-derived genotypes was then assigned using QUIRMIA- and EUCAST-based AST interpretation. QUIRMIA-based inference of resistance mechanisms and sequencing data were highly congruent (542/553, 98%). In contrast, EUCAST-based classification with its binary classification into "susceptible" and "resistant" isolates failed to recognize and properly categorize low-level resistant isolates. QUIRMIA provides a coherent genotype/phenotype categorization and may be integrated in the EUCAST expert rule set, thereby enabling reliable detection of low-level resistant isolates, which may help to better predict outcome and to prevent the emergence of clinical resistance.

Effects of the order of exposure to antimicrobials on the incidence of multidrug-resistant Pseudomonas aeruginosa

Multidrug-resistant Pseudomonas aeruginosa (MDRP) is one of the most important pathogens in clinical practice. To clarify the mechanisms contributing to its emergence, we isolated MDRPs using the P. aeruginosa PAO1, the whole genome sequence of which has already been elucidated. Mutant strains resistant to carbapenems, aminoglycosides, and new quinolones, which are used to treat P. aeruginosa infections, were isolated; however, none met the criteria for MDRPs. Then, PAO1 strains were exposed to these antimicrobial agents in various orders and the appearance rate of MDRP varied depending on the order of exposure; MDRPs more frequently appeared when gentamicin was applied before ciprofloxacin, but were rarely isolated when ciprofloxacin was applied first. Exposure to ciprofloxacin followed by gentamicin increased the expression of MexCD-OprJ, an RND-type multidrug efflux pump, due to the NfxB mutation. In contrast, exposure to gentamicin followed by ciprofloxacin resulted in more mutations in DNA gyrase. These results suggest that the type of quinolone resistance mechanism is related to the frequency of MDRP and that the risk of MDRP incidence is highly dependent on the order of exposure to gentamicin and ciprofloxacin.

Synergistic Effect of SOS Response and GATC Methylome Suppression on Antibiotic Stress Survival in Escherichia coli.

The suppression of the SOS response has been shown to enhance the in vitro activity of quinolones. Furthermore, Dam-dependent base methylation has an impact on susceptibility to other antimicrobials affecting DNA synthesis. Here, we investigated the interplay between these two processes, alone and in combination, in terms of antimicrobial activity. A genetic strategy was used employing single- and double-gene mutants for the SOS response (recA gene) and the Dam methylation system (dam gene) in isogenic models of Escherichia coli both susceptible and resistant to quinolones. Regarding the bacteriostatic activity of quinolones, a synergistic sensitization effect was observed when the Dam methylation system and the recA gene were suppressed. In terms of growth, after 24 h in the presence of quinolones, the Δdam ΔrecA double mutant showed no growth or delayed growth compared to the control strain. In bactericidal terms, spot tests showed that the Δdam ΔrecA double mutant was more sensitive than the ΔrecA single mutant (about 10- to 102-fold) and the wild type (about 103- to 104-fold) in both susceptible and resistant genetic backgrounds. Differences between the wild type and the Δdam ΔrecA double mutant were confirmed by time-kill assays. The suppression of both systems, in a strain with chromosomal mechanisms of quinolone resistance, prevents the evolution of resistance. This genetic and microbiological approach demonstrated the enhanced sensitization of E. coli to quinolones by dual targeting of the recA (SOS response) and Dam methylation system genes, even in a resistant strain model.

Proteus mirabilis isolated from untreated hospital wastewater, Ibadan, Southwestern Nigeria showed low-level resistance to fluoroquinolone and carried qnrD3 on Col3M plasmids.

Untreated wastewater emanating from healthcare facilities are risk factors for the spread of antimicrobial resistance (AMR) at the human-environment interface. In this study, we investigated the determinants of resistance in three multidrug resistant strains of Proteus mirabilis isolated from untreated wastewater collected from three government owned hospitals in Ibadan, Nigeria. Despite showing low-level resistance to ciprofloxacin, whole genome sequencing revealed the transferable mechanism of quinoloneresistance (TMQR)gene qnrD3 carried on Col3M plasmids in all the isolates. Core genome phylogenetic analysis showed the isolates are closely related differing from each other by ≤ 23 single nucleotide polymorphisms (SNP). Further, they shared the closest evolutionary relationship with isolates from China. Similarly, the Col3M plasmids is most closely related to p3M-2A found in P. vulgaris 3M isolated from the intestine of shrimps in China. This to the best of our knowledge is the first report of Col3M plasmids carrying qnrD3 in environmental bacterial isolates. Our results indicate a possible silent spread of this important plasmid associated with the dissemination of qnrD3 in Nigeria, and further highlights the important role played by untreated wastewater from healthcare facilities in the spread of AMR in low- and middle-income countries.

Prevalence and characterization of quinolone-resistant Escherichia coli isolated from retail raw beef and poultry meat in Egypt.

The goal of this study was to look at quinolone-resistant (QR) Escherichia coli (E. coli) from retail beef and poultry meat in Egypt by looking at the QR mechanisms in the resistant strains. In total, 120 samples of raw poultry meat (n = 60) and beef meat (n = 60) were purchased from Mansoura retail stores between January and March 2021, and evaluated microbiologically for E. coli. Then, an antimicrobial sensitivity test was applied to all isolates. The prevalence of QR E. coli with concern for the QR determinants, including quinolone resistance-determining regions (QRDRs) mutations, the plasmid-mediated quinolone resistance gene (PMQR), and the efflux pump activity were determined. The total prevalence of E. coli was 34.2% (41/120). Noticeably, the prevalence of E. coli in poultry meat (40%, 24/60) was higher than that of beef (28%, 17/60). All strains were assessed for their antimicrobial susceptibility using the disc diffusion technique; the highest rate of resistance (100%) was displayed to clindamycin and cefuroxime, followed by ampicillin (97.6%), doxycycline (92.7%), amoxicillin-clavulanate (92.7%), nalidixic acid (NA) (80.5%), sulfamethoxazole/trimethoprim (70.7%), chloramphenicol (63.4%), gentamicin, and azithromycin (58.5% each). Multiple antimicrobial resistance (strains resistant to three or more antimicrobial classes) was displayed by 97.6% of E. coli isolates. Regarding QR, 37 isolates could resist at least one of the examined quinolones. Regarding PMQR genes, qnrS was determined in 70% (7/10) of QR E. coli, while qnrA, qnrB, and qnrD were not identified. While the mutations determined regions of QR in the resistant E. coli isolates, S83L was the most prevalent in gyrase subunit A either alone or combined with D87N and D87Y, and three isolates of QR E. coli isolates revealed a topoisomerase IV subunit mutation harboring S80I. 20% of the isolates displayed efflux activity, as NA showed a considerable difference between its zones of inhibition. The high prevalence of antimicrobial-resistant E. coli, with concern for QR strains harboring different resistance mechanisms in poultry meat and beef, threatens the public's health. Thus, standard manufacturing procedures and adequate hygiene conditions must be followed in all phases of meat preparation, production, and consumption, and public knowledge should be improved.

Aquatic Environments as Hotspots of Transferable Low-Level Quinolone Resistance and Their Potential Contribution to High-Level Quinolone Resistance.

The disposal of antibiotics in the aquatic environment favors the selection of bacteria exhibiting antibiotic resistance mechanisms. Quinolones are bactericidal antimicrobials extensively used in both human and animal medicine. Some of the quinolone-resistance mechanisms are encoded by different bacterial genes, whereas others are the result of mutations in the enzymes on which those antibiotics act. The worldwide occurrence of quinolone resistance genes in aquatic environments has been widely reported, particularly in areas impacted by urban discharges. The most commonly reported quinolone resistance gene, qnr, encodes for the Qnr proteins that protect DNA gyrase and topoisomerase IV from quinolone activity. It is important to note that low-level resistance usually constitutes the first step in the development of high-level resistance, because bacteria carrying these genes have an adaptive advantage compared to the highly susceptible bacterial population in environments with low concentrations of this antimicrobial group. In addition, these genes can act additively with chromosomal mutations in the sequences of the target proteins of quinolones leading to high-level quinolone resistance. The occurrence of qnr genes in aquatic environments is most probably caused by the release of bacteria carrying these genes through anthropogenic pollution and maintained by the selective activity of antimicrobial residues discharged into these environments. This increase in the levels of quinolone resistance has consequences both in clinical settings and the wider aquatic environment, where there is an increased exposure risk to the general population, representing a significant threat to the efficacy of quinolone-based human and animal therapies. In this review the potential role of aquatic environments as reservoirs of the qnr genes, their activity in reducing the susceptibility to various quinolones, and the possible ways these genes contribute to the acquisition and spread of high-level resistance to quinolones will be discussed.

High frequency of non-susceptibility to ciprofloxacin in nontyphoid Salmonella recovered from Brazilian broiler chicken

ABSTRACT 1. This study evaluated the minimal inhibitory concentration (MIC) of ciprofloxacin and the presence of plasmid-mediated quinolone resistance (PMQR) mechanisms in 97 nontyphoidal Salmonella spp. isolated from broilers and carcases from three different regions in Brazil. The presence of mutations in quinolone resistance determination regions (QRDRS) was investigated in the ciprofloxacin-resistant strain by DNA sequencing. 2. Most of the Salmonella spp. (85.6%) had intermediate resistance to ciprofloxacin and only one isolate was resistant. MIC breakpoints ranged from ≤0.03 to 1 µg/ml and 67.0% of the strains had a MIC of 0.25 µg/ml (n=65). Thirteen strains (13.4%) were susceptible to ciprofloxacin with MIC ≤0.06 µg/ml. The qnrB gene was detected in eight isolates with intermediate resistance and in two susceptible strains. The other PMQR genes, qnrA, qnrC, qnrD, qnrS, qnrVC, aac(6′)-Ib, qepA, oqxAB and mutations in QRDR were not detected in any strain. 3. There was a high frequency of ciprofloxacin intermediate resistant Salmonella from broiler and broiler carcases from Brazil. The presence of these strains in poultry and derived products poses a risk to public health.

Impact on quinolone resistance of plasmid-mediated quinolone resistance gene and mutations in quinolone resistance-determining regions in extended spectrum beta lactamase-producing Klebsiella pneumoniae isolated from urinary tract infection patients.

Klebsiella pneumoniae is a typical pathogen in urinary tract infections (UTI), and the emergence of extended spectrum beta-lactamase (ESBL)-producing strains has been frequently reported, accompanied by higher quinolone resistance rates. There are two major mechanisms of quinolone resistance, mutations in quinolone resistance-determining regions (QRDR) and the presence of the plasmid-mediated quinolone resistance (PMQR) genes. This study aimed to investigate quinolone resistance among 105 ESBL-producing K. pneumoniae specimens isolated from UTI patients in Indonesia. These were characterized for antimicrobial resistance to nalidixic acid, ciprofloxacin, and levofloxacin, QRDR mutations in gyrA and parC and the presence of PMQR genes. We found that 84.8% of the collected isolates were resistant to at least one of the quinolones. QRDR mutation in gyrA was observed in 49.5% of these strains and parC mutations in 61.0%. PMQR genes were identified in 84.8% of strains. The QRDR mutations clearly had a greater effect on resistance than the PMQR genes. In conclusion, we found high quinolone resistance rates in Indonesian ESBL-producing K. pneumoniae, in which QRDR mutation played a major role.

The First Saudi Report of Novel and Common Mutations in the gyrA and parC Genes Among Pseudomonas Spp. Clinical Isolates Recovered from Taif Area.

Background and AimsReports examine quinolone resistance mechanisms among Pseudomonas spp. are sporadic in the Kingdom of Saudi Arabia (KSA). We previously examined the genetic bases of plasmid-mediated quinolone resistance among Pseudomonas spp. clinical isolates. This study investigated chromosomally mediated quinolone resistance mechanisms via investigation of the mutations in the gyrA and parC genes.MethodsThe minimum inhibitory concentration (MIC) to different quinolones was determined. Twenty-nine quinolone resistant Pseudomonas spp. clinical isolates were included. The gyrA and parC genes were sequenced by Sanger capillary electrophoresis. Multiple sequence alignment for the translated gyrA and parC genes was performed to identify mutation sites.ResultsOf the 29 isolates, 27 isolates were P. aeruginosa and two were P. putida. The cluster analysis of the quinolone susceptibility pattern revealed seven susceptibility phenotypes (A-G) based on susceptibility patterns rather than the MIC values. Also, 22 different susceptibility phenotypes were detected based on MIC values. All isolates exhibited a missense mutation at position 83 (S83I/T/F) of the gyrA gene in addition to six missense mutations at positions outside the QRDR of this gene. In addition, 82.8% (24/29) of the isolates harbored a missense mutation in the parC gene at position 87 (S87L), along with six novel mutations outside the QRDR of the parC gene. Haplotyping of the gyrA, parC, and the overall QRDR revealed six, 10, and 13 different haplotypes, respectively.ConclusionThis study documents the incidence of the commonly reported mutations in the gyrA and parC genes in addition to novel mutations in these genes among Pseudomonas spp. clinical isolates recovered from KSA. Together with our previous findings, these data provide an insight into the genetic background of quinolone resistance among Pseudomonas spp. clinical isolates in KSA.

Molecular Characterization of Quinolone Resistance Determinants in Non-Typhoidal Salmonella Strains Isolated in Tehran, Iran

Background: Quinolone resistant Salmonella serotypes have been reported in recent years and have become increasingly widespread worldwide. Objectives: We evaluated the molecular mechanism of quinolone resistance in non-typhoidal Salmonella strains isolated from clinical samples in Tehran, Iran. Methods: The present study included the Salmonella isolates originated from hospitalized individuals and outpatients in Tehran, Iran. Serotyping of nalidixic acid-resistant Salmonella isolates was done by slide agglutination method. Then, the quinolone resistance-determining region (QRDR) of topoisomerase gene gyrA and the plasmid-mediated quinolone resistance (PMQR) determinants were detected using the polymerase chain reaction (PCR) method. Restriction fragment length polymorphism (RFLP) analysis was also employed to determine the possible mutation in the gyrA gene of those strains. Mutant strains were detected by enzymatic digestion, and their PCR products were sequenced immediately. Results: Amongst 141 isolates, 60% showed nalidixic acid resistance, whereas none of them were ciprofloxacin-resistant. The commonly prevalent serotypes were S. Enteritidis and S. Infantis. Of 85 nalidixic acid-resistant strains, 17 (20%) isolates harbored the qnrS gene. However, PCR analysis of the quinolone-resistant strains did not detect qnrA and qnrB genes. PCR-RFLP and sequencing analysis of the QRDRs of the gyrA gene indicated that 16 (18.8%) isolates had mutant patterns, and the most common point mutation was serine to phenylalanine at position 83. Conclusions: Our results demonstrated that point mutations in gyrA and the existence of plasmid-mediated gene qnrS were important mechanisms of quinolone resistance in non-typhoidal Salmonella strains isolated from human origin. Other alternative mechanisms of resistance, such as alterations in the expression of efflux pumps, should be studied to provide greater insight into the molecular mechanism of quinolone-resistant non-typhoidal Salmonella isolates.

Epidemiology of Multidrug Resistance among Salmonella enterica serovars typhi and paratyphi A at a Tertiary Pediatric Hospital in India Over a Decade; In-silico Approach to Elucidate the Molecular Mechanism of Quinolone Resistance

BackgroundEnteric fever caused by Salmonella enterica serovar typhi and Salmonella enterica serovar paratyphi A remains one of the most common causes of community-acquired bloodstream infection among children in India. Multidrug resistance is emerging and is a cause of concern as it affects the choice of treatment in enteric fever. MethodIn this study, a 10-year analysis of resistance patterns was done along with in-silico homology modeling and molecular docking to understand the commonly occurring quinolone resistance. ResultsA total of 1010 cases of blood culture–confirmed enteric fevers (S. typhi n=849; S. paratyphi A n=161) were recorded at the study hospital during the period from 2011-2020. Multidrug resistance among cases of S. typhi was 2.12 %, whereas it was completely absent among cases of S. paratyphi A. Fluoroquinolone resistance was high (>95%) throughout the study period. Resistance to ampicillin, chloramphenicol and co-trimoxacole was low (<3%) among S. typhi cases. No deaths were observed among study participants. Molecular docking analysis showed that quinolone had less binding affinity to mutated gyrase A than to its wild type for both S. typhi and S. paratyphi A. ConclusionQuinolone resistance was high among cases of enteric fever, whereas no resistance was observed among third-generation cephalosporins. In-silico studies indicated that a mutation in gyrase A might be the cause of the gradual increase in ciprofloxacin resistance over the study period.