Fluoroquinolone-resistant Isolates Research Articles

Longitudinal molecular analysis of clinical and fecal Escherichia coli isolates at a Veterans Affairs Medical Center in Minnesota, USA, 2012-2019.

Extraintestinal Escherichia coli infections represent a growing public health threat, However, current studies often overlook important factors such as temporal patterns of infection, phylogenetic and clonal background, or the host gut E. coli population, despite their likely significance. In this study, we analyzed >7000 clinical E. coli isolates from patients at the Minneapolis Veterans Affairs Health Care System (2012-2019), and concurrent fecal E. coli from uninfected veterans. We assessed phylogenetic group distribution, membership in selected sequence types (STs), and subsets thereof-including the pandemic, resistance-associated ST131-H30R, and ST1193 lineages-and strain type, as defined by pulsed-field gel electrophoresis. We then analyzed these features alongside the temporal patterns of infection in individual hosts. The H30R lineage emerged as the leading lineage, both overall and among fluoroquinolone-resistant isolates, with ST1193 following among fluoroquinolone-resistant isolates. Recurrences were common, occurring in 31% of subjects and 41% of episodes, and often multiple and delayed/prolonged (up to 23 episodes per subject; up to 2655d post-index). Remarkably, these recurrences typically involved the subject's index strain (63% of recurrences), even when affecting extra-urinary sites. ST131, H30R, ST1193, and fluoroquinolone-resistant strains generally caused significantly more recurrences than did other strains, despite similar recurrence intervals. ST131 strain types shifted significantly over the study period. Infection-causing strains were commonly detectable in host feces at times other than during an infection episode; the likelihood of detection varied with surveillance intensity and proximity to the infection. H30R and ST1193 were prominent causes of fecal-clinical clonal overlap. These findings provide novel insights into the temporal and clonal characteristics of E. coli infections in veterans and support efforts to develop anti-colonization interventions.

Coexistence of plasmid-mediated quinolone resistance (PMQR) and extended-spectrum beta-lactamase (ESBL) genes among clinical Pseudomonas aeruginosa isolates in Egypt

BackgroundData about the prevalence of plasmid-mediated quinolone resistance (PMQR) and extended-spectrum beta-lactamase (ESBL) production in P. aeruginosa compared to the Enterobacteriaceae family is limited. The availability of limited therapeutic options raises alarming concerns about the treatment of multidrug-resistant P. aeruginosa. This study aimed to assess the presence of PMQR and ESBL genes among P. aeruginosa strains.MethodsFifty-six P. aeruginosa strains were isolated from 330 patients with different clinical infections. Phenotypically fluoroquinolone-resistant isolates were tested by PCR for the presence of six PMQR genes. Then, blaTEM, blaSHV, and blaCTX-M type ESBL genes were screened to study the co-existence of different resistance determinants.ResultsOverall, 22/56 (39.3%) of the studied P. aeruginosa isolates were phenotypically resistant to fluoroquinolones. PMQR-producing P. aeruginosa isolates were identified in 20 isolates (90.9%). The acc(6')-Ib-cr was the most prevalent PMQR gene (77.3%). The qnr genes occurred in 72.7%, with the predominance of the qnrA gene at 54.5%, followed by the qnrS gene at 27.3%, then qnrB and qnrC at 22.7%. The qepA was not detected in any isolate. The acc(6')-Ib-cr was associated with qnr genes in 65% of positive PMQR isolates. Significant differences between the fluoroquinolone-resistant and fluoroquinolone-susceptible isolates in terms of the antibiotic resistance rates of amikacin, imipenem, and cefepime (P value < 0.0001) were found. The ESBL genes were detected in 52% of cephalosporin-resistant P. aeruginosa isolates. The most frequent ESBL gene was blaCTX-M (76.9%), followed by blaTEM (46.2%). No isolates carried the blaSHV gene. The acc(6')-Ib-cr gene showed the highest association with ESBL genes, followed by the qnrA gene. The correlation matrix of the detected PMQR and ESBL genes indicated overall positive correlations. The strongest and most highly significant correlation was between qnrA and acc(6')-Ib-cr (r = 0.602) and between qnrA and blaCTX-M (r = 0.519).ConclusionA high prevalence of PMQR genes among the phenotypic fluoroquinolone-resistant P. aeruginosa isolates was detected, with the co-carriage of different PMQR genes. The most frequent PMQR was the acc(6')-Ib-cr gene. Co-existence between PMQR and ESBL genes was found, with 75% of PMQR-positive isolates carrying at least one ESBL gene. A high and significant correlation between the ESBL and PMQR genes was detected.

Identification of novel resistance-associated mutations and discrimination within whole-genome sequences of fluoroquinolone-resistant Mycobacterium tuberculosis isolates.

This study aims to elucidate additional mutation loci associated with fluoroquinolone (FQ) resistance and evaluate the discriminatory capacity of mutation loci and allele mutation frequencies in identifying FQ-resistant Mycobacterium tuberculosis (MTB) isolates. A random selection of isolates was extracted from an ongoing collection. Drug resistance was determined using the resazurin microtiter assay (REMA) as the gold standard. Mutation loci and the burden of mutations in the quinolone resistance-determining region (QRDR) were elucidated through whole-genome sequencing (WGS). Novel amino acid mutations, namely, G520D and G520T, were identified in the gyrB and associated with FQ resistance. In the context of distinguishing FQ-resistant isolates, the AUC for the QRDR mutation frequency burden (0.969) surpassed that of the mutation locus (0.929), and this difference was statistically significant (P = 0.03). Furthermore, using the resistance mutation locus as a reference, setting the QRDR mutation frequency burden threshold at 1.31% resulted in a 3.60% increase in the accuracy of classifying FQ-resistant isolates (NRI = 3.60%, P < 0.001). The QRDR mutation frequency burden appears to offer superior diagnostic efficacy in discriminating FQ-resistant isolates compared to qualitative detection of mutant loci.IMPORTANCEFluoroquinolone (FQ) drugs are recommended as second-line drugs for the treatment of multidrug-resistant tuberculosis. With the massive use of FQ drugs in the clinical treatment of tuberculosis (TB), there is an increasing rate of drug resistance to FQ drugs. In this study, we identified and demonstrated novel amino acid mutations associated with FQ resistance in Mycobacterium tuberculosis (MTB), and we quantified the mutation sites and identified the quinolone resistance-determining region (QRDR) mutation frequency burden as a novel diagnostic method for FQ resistance. We hope that the results of this study will provide data support and a theoretical basis for the rapid diagnosis of FQ-resistant MTB.

Virulence genes, resistome and mobilome of Streptococcus suis strains isolated in France.

Streptococcus suis is a leading cause of infection in pigs, causing extensive economic losses. In addition, it can also infect wild fauna, and can be responsible for severe infections in humans. Increasing antimicrobial resistance (AMR) has been described in S. suis worldwide and most of the AMR genes are carried by mobile genetic elements (MGEs). This contributes to their dissemination by horizontal gene transfer. A collection of 102 strains isolated from humans, pigs and wild boars in France was subjected to whole genome sequencing in order to: (i) study their genetic diversity, (ii) evaluate their content in virulence-associated genes, (iii) decipher the mechanisms responsible for their AMR and their association with MGEs, and (iv) study their ability to acquire extracellular DNA by natural transformation. Analysis by hierarchical clustering on principal components identified a few virulence-associated factors that distinguish invasive CC1 strains from the other strains. A plethora of AMR genes (n=217) was found in the genomes. Apart from the frequently reported erm(B) and tet(O) genes, more recently described AMR genes were identified [vga(F)/sprA, vat(D)]. Modifications in PBPs/MraY and GyrA/ParC were detected in the penicillin- and fluoroquinolone-resistant isolates respectively. New AMR gene-MGE associations were detected. The majority of the strains have the full set of genes required for competence, i.e for the acquisition of extracellular DNA (that could carry AMR genes) by natural transformation. Hence the risk of dissemination of these AMR genes should not be neglected.

Prevalence and molecular characterization of multi-resistant Escherichia coli isolates from clinical bovine mastitis in China

Different antibiotics are used to treat mastitis in dairy cows that is caused by Escherichia coli (E. coli). Antimicrobial resistance in food-producing animals in China has been monitored since 2000. Surveillance data have shown that the prevalence of multiresistant E. coli in animals has increased significantly. This study aimed to investigate the occurrence and molecular characteristics of resistance determinants in E. coli strains (n = 105) obtained from lactating cows with clinical bovine mastitis (CBM) in China. A total of 220 cows with clinical mastitis, which has swollen mammary udder with reduced and red or gangrenous milk, were selected from 5000 cows. The results showed 94.3% of the isolates were recognized as multidrug resistant. The isolates (30.5%) were positive for the class I integrase gene along with seven gene cassettes that were accountable for resistance to trimethoprim resistance (dfrA17, dfr2d and dfrA1), aminoglycosides resistance (aadA1 and aadA5) and chloramphenicol resistance (catB3 and catB2), respectively. The bla TEM gene was present in all the isolates, and these carried the bla CTX gene. A double mutation in gyrA (i.e., Ser83Leu and Asp87Asn) was observed in all fluoroquinolone-resistant isolates. In total, nine fluoroquinolone-resistant E. coli isolates were identified with five different types of mutations in parC. In four (44.4%) isolates, Ser458Ala was present in parE, and in all nine (9/9) fluoroquinolone-resistant isolates, Pro385Ala was present in gyrB. Meanwhile, fluoroquinolone was observed as highly resistant, especially in isolates with gyrA and parC mutations. In summary, the findings of this research recognize the fluoroquinolone resistance mechanism and disclose integron prevalence and ESBLs in E. coli isolates from lactating cattle with CBM.

High Genetic Diversity in Third-Generation Cephalosporin-Resistant Escherichia coli in Wastewater Systems of Schleswig-Holstein.

The spread of multidrug-resistant bacteria from humans or livestock is a critical issue. However, the epidemiology of resistant pathogens across wastewater pathways is poorly understood. Therefore, we performed a detailed comparison of third-generation cephalosporin-resistant Escherichia coli (3GCREC) from wastewater treatment plants (WWTPs) to analyze dissemination pathways. A total of 172 3GCREC isolated from four WWTPs were characterized via whole genome sequencing. Clonal relatedness was determined using multi-locus sequence typing (MLST) and core genome MLST. Resistance genotypes and plasmid replicons were determined. A total of 68 MLST sequence types were observed with 28 closely related clusters. Resistance genes to eight antibiotic classes were detected. In fluoroquinolone-resistant isolates, resistance was associated with three-or-more point mutations in target genes. Typing revealed high genetic diversity with only a few clonal lineages present in all WWTPs. The distribution paths of individual lines could only be traced in exceptional cases with a lack of enrichment of certain lineages. Varying resistance genes and plasmids, as well as fluoroquinolone resistance-associated point mutations in individual isolates, further corroborated the high diversity of 3GCREC in WWTPs. In total, we observed high diversity of 3GCREC inside the tested WWTPs with proof of resistant strains being released into the environment even after treatment processes.

Prevalence of Brucella melitensis and Brucella abortus Fluoroquinolones Resistant Isolates: A Systematic Review and Meta-Analysis.

Background: Brucellosis impact both animals and humans worldwide. However, using antibiotics for brucellosis remains controversial despite decades of research. Relapse can complicate treatment in this area. Since the mid-1980s, microbiologists, and physicians have studied fluoroquinolones' use for treating human brucellosis. The principal advantages of fluoroquinolones are their intracellular antimicrobial activity, low nephrotoxicity, good pharmacokinetics, and the lack of drug-level monitoring. Fluoroquinolones inhibit disease recurrence. In vitro and clinical data were used to study the prevalence of Brucella melitensis and Brucella abortus fluoroquinolone-resistant isolates. Methods: The PubMed, Scopus, Embase, and Web of Science databases were carefully searched until August 6, 2022, for relevant papers. The number of resistant isolates and sample size were used to estimate the proportion of resistant isolates, fitting a model with random effects, and DerSimonian-Laird estimated heterogeneity. Furthermore, meta-regression and subgroup analyses were used to assess the moderators to identify the sources of heterogeneity. Meta-analysis was performed using R software. Results: Forty-seven studies evaluated fluoroquinolone resistance in Brucella spp. Isolates. Fluoroquinolones have shown high in vitro efficacy against Brucella spp. The resistance rates to ofloxacin, sparfloxacin, fleroxacin, pefloxacin, and lomefloxacin were 2%, 1.6%, and 4.6%, respectively. Conclusion: Clinical in vitro tests demonstrated that fluoroquinolones can eradicate Brucella spp. Owing to first-line medication resistance, recurrence, and toxicity, it is essential to standardize the Brucella antimicrobial susceptibility test method for a more precise screening of resistance status. Fluoroquinolones are less resistant to fluoroquinolone-based treatments in modern clinical practice as alternatives to standard therapy for patients with brucellosis relapse after treatment with another regimen and in patients who have developed toxicity from older agents.

Antimicrobial susceptibility and molecular characteristics of beta-lactam- and fluoroquinolone-resistant E. coli from human clinical samples in Nigeria

This study determined the antimicrobial resistance profiles and molecularly characterized beta-lactam- and fluoroquinolone-resistant E. coli recovered from human clinical samples. Three hundred (300) clinical samples collected from two major tertiary healthcare institutions were analyzed using standard microbiological techniques. Antimicrobial susceptibility testing and phenotypic detection of extended-spectrum beta-lactamase (ESBL) were done by disc diffusion and Double Disk Synergy Test. Molecular characterization for ESBL and fluoroquinolone-resistant genes were done by PCR with specific primers. In total, 44 (14.7%) ESBL-producing E. coli were recovered from human clinical specimens. The recovered isolates were multidrug-resistant with multiple antibiotics resistance index values ranging from 0.5–0.8. Isolates exhibited resistance to sulfamethoxazole/trimethoprim 44 (100%), cefoxitin 42 (95.5%), tetracycline 42 (95.5 %), cefuroxime 41 (93.2%), ciprofloxacin 40 (90.9%), cefotaxime 37 (84.1%), amoxicillin 37 (84.1%), amoxicillin/clavulanic acid 37 (84.1), and ceftazidime 35 (79.5%). BlaTEM 41 (93.2%) ESBL gene was the most predominant gene among the isolates, followed by blaCTX-M 9 (20.5%), and blaSHV 1 (2.3%) genes. Co-existence of blaTEM +blaSHV, blaTEM + blaCTX-M, and blaTEM +blaSHV + blaCTX-M genes was observed in 1 (2.3%), 7 (15.9%), and 4 (9.1%) isolates, respectively. All the fluoroquinolone-resistant isolates harbored aac-lb-6-cr gene while Qnr gene was absent. Association of fluoroquinolone resistance- and β-lactam resistance genes; aac-lb-6-cr + blaTEM + blaSHV + blaCTM was observed in 5 (11.4%) isolates. Our study showed high frequency of ESBL-producing E. coli harboring ESBL and fluoroquinolone resistance genes from human clinical samples in Ebonyi State, Nigeria. The high frequency of multidrug-resistant E. coli in our study is very worrisome and could have significant public health impact such as treatment failures, and possibly death, if not properly managed. It is therefore imperative to establish strong regulative measures and guidelines that would help in curtailing the increasing dissemination of these superbugs in healthcare institutions in Nigeria.

Antibiotic Resistance Pattern and Distribution of Resistance Genes in Salmonella Isolated from Chicken and Duck Eggs in Chhattisgarh, India

Background: Salmonella is recognized as the most prevalent bacterial cause of foodborne diseases worldwide and animal-sourced foods have been reported as a common source of Salmonella infections among humans. Methods: The commercial chicken eggs, backyard chicken eggs, and duck eggs samples, 60 each, were processed for isolation and identification of Salmonella. All Salmonella isolates were further tested for resistance against six different antibiotics. The prevalence of virulence and antimicrobial resistance genes in the Salmonella isolates was determined by PCR. Result: A total of 28 Salmonella isolates were recovered with an overall prevalence of 15.6% and out of them, 11.1% and 4.4% were from eggshell and egg content, respectively. All the isolates were found sensitive to Gentamicin however maximum resistance was observed against Cefotaxime. PCR results revealed that 100% of the isolates were carrying the invA gene however stn gene was detected in 78.6% of isolates. Among presumptively identified β-lactam-resistant Salmonella isolates, 100% and 50% isolates harbored blaTEM and blaCTX-M genes, respectively whereas none of the isolates contained the blaSHV gene. All tetracycline-resistant isolates harbored the tetA gene whereas none of the isolates carried the tetB gene. 100% of fluoroquinolone-resistant isolates were carrying the gyrA gene however parC gene was present only in 60% of isolates. These results indicate that drug-resistant Salmonella spp. were prevalent in eggs sold in the study area which can pose a serious public health problem.

Pattern and characteristics of mutations conferring resistance to second line drugs in Mycobacterium tuberculosis isolates of pulmonary and extrapulmonary TB samples

Background & objectivesThe purpose of present study is to analyse the distribution and pattern of genetic mutations in PRE-XDR-TB and extensive drug resistant Mycobacterium tuberculosis (XDR-TB) using second-line line probe assay and to compare them with different parameters. MethodSputum, Lymph node aspirate and cold accesses from patients with rifampicin resistant Tuberculosis were subjected to first line and second line Probe Assay (Genotype MTBDRsl by Hain Life Science, Germany) to assess additional drug resistance to fluroquinolones (Levofloxacin & Moxifloxacin) and Aminoglycosides (Amikacin, Ofloxacin and Kanamycin). The genetic mutation pattern was analysed and compared with demographic, clinical and other parameters. ResultsThe final study population included 123 fluoroquinolone resistant isolates including 14 isolates with additional second line aminoglycosides drug resistance. The most frequent mutation observed among Gyr A drug resistance mutation was D94G (Gyr A MUT3C, 50/123,40%) corresponding to high level resistance to levofloxacin and moxifloxacin. The most frequent wild type mutant among Gyr A gene locus was WT 3 (85/123,69%). The most common mutation among second line aminoglycoside resistant isolates was at eis WT2 (7/14,50%) followed by rrs MUT 2 (4/14,29%). ConclusionsGyrA MUT3C (Asp94Gly) was the most common mutation in Gyr A gene locus in M. tuberculosis causing high level levofloxacin and moxifloxacin resistance. Patients with Asp94Gly mutation was significantly associated with underweight body mass index (p = 0.026). This study also observed that history of anti-tuberculosis therapy is a risk factor for FQ drug resistance mutations (p < 0.001).

Analysing the Impact of Bedaquiline Resistance in Fluoroquinolone-Resistant Clinical Isolates and Investigating the Molecular Interactions of BDQ with the atpE Gene

The progress of the tuberculosis eradication programme is hindered by the global health issue of drug-resistant tuberculosis, which is a significant threat to the population. Resistance-associated mutations vary geographically, so investigate the prevalence of resistance to fluoroquinolone (anti-TB) drugs and its association with resistance-related mutations in clinical isolates of Tamilnadu (north zone) and Puducherry was carried out. And the resultant fluoroquinolone drug-resistant tuberculosis strains will be subjected to the molecular analysis of bedaquiline resistance to study the mutation in the atpE gene. The study includes 430 specimens received at Intermediate Reference Laboratory, State TB training and Demonstration Centre in the Government Hospital for Chest Diseases for Molecular testing between January 2020 and December 2021. Samples were analysed by GenoType MTBDRslV.2 for the molecular detection of mutations in the gyrA and gyrB genes. FQ resistance was observed in 32 samples (7.4%), most of which were naïve and previous treatment failure cases. Twenty-five isolates had mutations in DNA gyrase subunit -A (gyrA), while mutations in gyrB were observed in only 7 isolates. Mutational analysis revealed that the mutations mainly alter codon 94 (replacing aspartic acid with glycine, D94G), and 90 (replacing alanine with valine, A90V). In Isoniazid Resistance, MDR, and treatment failure cases, the resistance to Fluoroquinolones was most commonly associated with the D94G mutation. A molecular stimulation study evaluates the effect of BDQ on atpE gene by docking, using AutoDock 4.2, showing hydrogen, hydrophobic and electrostatic interactions.

Genomic insight into Campylobacter jejuni isolated from commercial turkey flocks in Germany using whole-genome sequencing analysis.

Campylobacter (C.) jejuni is a zoonotic bacterium of public health significance. The present investigation was designed to assess the epidemiology and genetic heterogeneity of C. jejuni recovered from commercial turkey farms in Germany using whole-genome sequencing. The Illumina MiSeq® technology was used to sequence 66 C. jejuni isolates obtained between 2010 and 2011 from commercial meat turkey flocks located in ten German federal states. Phenotypic antimicrobial resistance was determined. Phylogeny, resistome, plasmidome and virulome profiles were analyzed using whole-genome sequencing data. Genetic resistance markers were identified with bioinformatics tools (AMRFinder, ResFinder, NCBI and ABRicate) and compared with the phenotypic antimicrobial resistance. The isolates were assigned to 28 different sequence types and 11 clonal complexes. The average pairwise single nucleotide-polymorphisms distance of 14,585 SNPs (range: 0-26,540 SNPs) revealed a high genetic distinction between the isolates. Thirteen virulence-associated genes were identified in C. jejuni isolates. Most of the isolates harbored the genes flaA (83.3%) and flaB (78.8%). The wlaN gene associated with the Guillain-Barré syndrome was detected in nine (13.6%) isolates. The genes for resistance to ampicillin (bla OXA), tetracycline [tet(O)], neomycin [aph(3')-IIIa], streptomycin (aadE) and streptothricin (sat4) were detected in isolated C. jejuni using WGS. A gene cluster comprising the genes sat4, aph(3')-IIIa and aadE was present in six isolates. The single point mutation T86I in the housekeeping gene gyrA conferring resistance to quinolones was retrieved in 93.6% of phenotypically fluoroquinolone-resistant isolates. Five phenotypically erythromycin-susceptible isolates carried the mutation A103V in the gene for the ribosomal protein L22 inferring macrolide resistance. An assortment of 13 β-lactam resistance genes (bla OXA variants) was detected in 58 C. jejuni isolates. Out of 66 sequenced isolates, 28 (42.4%) carried plasmid-borne contigs. Six isolates harbored a pTet-like plasmid-borne contig which carries the tet(O) gene. This study emphasized the potential of whole-genome sequencing to ameliorate the routine surveillance of C. jejuni. Whole-genome sequencing can predict antimicrobial resistance with a high degree of accuracy. However, resistance gene databases need curation and updates to revoke inaccuracy when using WGS-based analysis pipelines for AMR detection.

Possible step-up in prevalence for Escherichia coli ST131 from fecal to clinical isolates: inferred virulence potential comparative studies within phylogenetic group B2

BackgroundEscherichia coli sequence type (ST)131 is an important urinary tract pathogen, and is responsible for considerable healthcare-associated problems and costs worldwide. A better understanding of the factors that contribute to its rapid worldwide spread may help in arresting its continual spread. We studied a large collection of fecal and urinary E. coli ST131 and E. coli non-ST131 phylogenetic group B2 isolates, from women, men and children, in regional NSW, Australia.ResultsWe found out that there was a step up in ST131 prevalence (and possibly in virulence) from fecal to clinical (urinary) isolates in general, and specifically among ciprofloxacin resistant isolates, in the 3 host groups. Furthermore, our results revealed that the inferred virulence potential of the ST131 isolates (as measured by VF gene scores) was much higher than that of non-ST131 phylogenetic group B2 isolates, and this was much more pronounced amongst the urinary isolates. This finding suggests presence of possible E. coli phylogenetic B2 subgroups with varying levels of virulence, with ST131 being much more virulent compared to others. A strong association between ST131 and fluoroquinolone (FQ) resistance was also demonstrated, suggesting that FQ use is related to ST131 emergence and spread. Specifically, about 77% of ST131 isolates from women and men, and 47% from children, were extended spectrum β- lactamase (ESBL) producers. Moreover, FQ resistant ST131 ESBL isolates on average harbored more VF genes than all other isolates.ConclusionsThe strong association between ST131 prevalence and FQ resistance amongst the studied isolates suggests that FQ use is related to ST131 emergence and spread. Furthermore, our results demonstrate that FQ resistance and a plurality of VF genes can exist together in ST131, something that has traditionally been regarded as being inversely related. This may partly contribute to the emergence and worldwide spread of ST131.

Phenotypic and genotypic antimicrobial susceptibility patterns of the emerging human respiratory pathogen Mycoplasma amphoriforme isolated from the UK and Denmark.

To determine the phenotypic and genotypic antibiotic susceptibility of Mycoplasma amphoriforme isolates recovered from patients in the UK and Denmark. Seven isolates of M. amphoriforme were examined for antimicrobial susceptibility to seven antibiotics using the microbroth dilution assay in line with the CLSI guidelines for mycoplasmas. Each isolate was additionally subjected to WGS to identify resistance-associated mutations. Based on the consensus sequences from the genomic data, PCR primers were designed, and tested, for the amplification of the QRDR within the parC gene. Of the seven isolates investigated, four (57%) were resistant to moxifloxacin (0.5-1 mg/L) and levofloxacin (1-2 mg/L), compared with those that were susceptible (0.03-0.06 and 0.006 mg/L, respectively). Isolate H29 was resistant to five of the seven antibiotics tested: moxifloxacin, 0.5 mg/L; levofloxacin, 2 mg/L; azithromycin, 64 mg/L; erythromycin, 128 mg/L; and clindamycin, 64 mg/L. All isolates were susceptible to tetracycline (0.06 mg/L) and lefamulin (0.001-0.004 mg/L). Mutations from genomic data confirmed the presence of an S89F mutation within the ParC protein among all fluoroquinolone-resistant isolates and an A2059G mutation in the 23S rRNA gene in the macrolide- and lincosamide-resistant isolate H29. To the best of our knowledge, this is the first time where phenotypic and genotypic resistance data have been paired for M. amphoriforme confirming a correlation between the two. These data suggest the need for focused testing and resistance determination of isolates from high-risk patients given the backdrop of a high prevalence of antimicrobial resistance.

Antimicrobial susceptibility profiles and tentative epidemiological cutoff values of Legionella pneumophila from environmental water and soil sources in China.

Legionnaires’ disease (LD), caused by Legionella, including the most prevalent Legionella pneumophila, has been treated primarily with antibiotics. Environmental water and soil are the reservoirs for L. pneumophila. Studying antimicrobial susceptibility using a large number of isolates from various environmental sources and regions could provide an unbiased result. In the present study, antimicrobial susceptibility of 1464 environmental L. pneumophila isolates that were derived from various environmental water and soil sources of 12 cities in China to rifampin (RIF), erythromycin (ERY), clarithromycin (CLA), azithromycin (AZI), ciprofloxacin (CIP), moxifloxacin (MOX), levofloxacin (LEV), and doxycycline (DOX) was investigated, and minimum inhibitory concentration (MIC) data were obtained. We show that regarding macrolides, ERY was least active (MIC90 = 0.5 mg/L), while CLA was most active (MIC90 = 0.063 mg/L). A total of three fluoroquinolones have similar MICs on L. pneumophila. Among these antimicrobials, RIF was the most active agent, while DOX was the most inactive one. We observed different susceptibility profiles between serogroup 1 (sg1) and sg2-15 or between water and soil isolates from different regions. The ECOFFs were ERY and AZI (0.5 mg/L), RIF (0.002 mg/L), CIP, CLA and MOX (0.125 mg/L), LEV (0.063 mg/), and DOX (32 mg/L). Overall, two fluoroquinolone-resistant environmental isolates (0.14%) were first documented based on the wild-type MIC distribution. Not all azithromycin-resistant isolates (44/46, 95.65%) harbored the lpeAB efflux pump. The MICs of the ERY and CLA on the lpeAB + isolates were not elevated. These results suggested that the lpeAB efflux pump might be only responsible for AZI resistance, and undiscovered AZI-specific resistant mechanisms exist in L. pneumophila. Based on the big MIC data obtained in the present study, the same defense strategies, particularly against both CLA and RIF, may exist in L. pneumophila. The results determined in our study will guide further research on antimicrobial resistance mechanisms of L. pneumophila and could be used as a reference for setting clinical breakpoints and discovering antimicrobial-resistant isolates in the clinic, contributing to the antibiotic choice in the treatment of LD.

Changes in the characteristics of community-onset fluoroquinolone-resistant Escherichia coli isolates causing community-acquired acute pyelonephritis in South Korea.

This study aimed to examine the changes in the characteristics of community-onset fluoroquinolone-resistant (FQ-R) Escherichia coli isolates causing community-acquired acute pyelonephritis (APN) in South Korea. Blood or urine samples were prospectively collected from patients aged ≥15 years with community-acquired APN who were admitted to one of the eight Korean hospitals included in this study between September 2017 and August 2018. Phylogenetic typing, multilocus sequence typing, and molecular characterization of β-lactamase resistance and plasmid-mediated quinolone resistance (PMQR) determinants were performed. The data were compared with those from a previous study with the same design conducted in 2010-2011. A total of 300 and 346 isolates were identified in 2010-2011 and 2017-2018, respectively. Among them, 76 (22.0%) and 77 (25.7%) FQ-R isolates were identified in 2010-2011 and 2017-2018, respectively. A significantly higher antimicrobial resistance against third-to fourth-generation cephalosporins, including cefotaxime (23.9% vs. 77.9%, P<0.001), were observed among FQ-R isolates in 2017-2018 than among those in 2010-2011. A higher proportion of ST131 isolates (27.6% vs. 66.2%, P<0.001), as well as isolates that had extended-spectrum β-lactamase (ESBL)/plasmid-mediated AmpC β-lactamase (PABL) (23.7% vs. 79.2%, P<0.001), was observed in 2017-2018 than in 2010-2011. Further, more PMQR determinants (11.8% vs. 40.8%, P<0.001) were observed in 2017-2018 than in 2010-2011. Among uropathogenic FQ-R E. coli isolates in South Korea, the prevalence of ST131 and the proportion of isolates containing ESBL and/or PMQR determinants have increased.

Clonal distribution and antimicrobial resistance of methicillin-susceptible and -resistant Staphylococcus aureus strains isolated from broiler farms, slaughterhouses, and retail chicken meat

Colonization of food-producing animals by antimicrobial-resistant Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA), has become a serious public health problem worldwide. In the current study, clonal diversities of livestock-associated S. aureus isolates collected from broiler farms, slaughterhouses, and retail chicken meat were examined. Two-hundred S. aureus isolates (43 MRSA and 157 methicillin-susceptible S. aureus [MSSA] isolates) were analyzed to determine 1) the genotypes of the isolates (multilocus sequence, agr, and spa types), 2) the methicillin resistance phenotype and staphylococcal cassette chromosome mec (SCCmec) types, 3) the antimicrobial resistance profiles, and 4) the mutational changes in gyrA, gyrB, parC, and parE in fluoroquinolone-resistant isolates. Fifteen different sequence types (STs) of MSSA strains displaying a relatively high degree of genetic diversity were detected in broiler farms, slaughterhouses, and retail chicken meat. In contrast to MSSA, 2 dominant genetic lineages of MRSA (ST692-SCCmecV with t2249 spa type, and ST188-SCCmecIVa with spa type t189) were found in healthy broilers. The high prevalence of ST692 and ST188 in healthy broilers is associated with high levels of multiple antimicrobial-resistance phenotypes, particularly fluoroquinolone resistance. All fluoroquinolone-resistant isolates carried double point mutations in gyrA (S84L) and parC (S80F), regardless of STs or methicillin resistance. Notably, only the ST188 lineage carried an additional third mutation in gyrB (D494N), correlating with enhanced ciprofloxacin minimum inhibitory concentration values versus the strains with double mutations. These results provide important insights into the genetic diversity of antimicrobial-resistant S. aureus strains associated with the chicken meat production chain, including healthy broilers, in Korea.

Ocular surface flora and prophylactic antibiotics for cataract surgery in the age of antimicrobial resistance.

According to the World Health Organization alert about the antimicrobial resistance crisis released in 2015, clinicians should strongly reconsider the prolonged use of antimicrobials. In this review, we focus on the ocular surface flora with respect to the trend of fluoroquinolone resistance, and its upset and restoration after topical administration of antimicrobials and preservatives. Even 3weeks of topical administration of levofloxacin (LVFX) yields a selection of fluoroquinolone-resistant isolates bearing genetic changes in the ocular surface flora. One month of topical prophylactic administration of LVFX after cataract surgery induces the loss of diversity with LVFX-resistance of the ocular surface flora. Restoration of LVFX-sensitive flora occurs 6 to 9months after the final topical administration of LVFX. The ocular surface flora recovers earlier in patients given LVFX for 1week after the surgical procedure. These findings suggest that shorter periods of postoperative topical antibiotics are less frequently associated with persistent antimicrobial-resistant bacteria in the ocular flora. In addition, microbiologic analysis of ocular surfaces treated with a long period of eye drops containing benzalkonium chloride (BAC) showed a higher incidence of isolates resistant to methicillin and fluoroquinolones than did ocular surfaces treated with eye drops not containing BAC. To avoid the emergence of antimicrobial-resistant bacteria on the ocular surface, an urgent discussion must be held about the appropriate use of antibiotics and preservatives in the ophthalmology field.

A survey of antimicrobial-resistant Escherichia coli prevalence in wild mammals in Japan using antimicrobial-containing media.

The emergence and spread of antimicrobial-resistant bacteria and resistance genes pose serious human and animal health concerns. Therefore, to control antimicrobial-resistant bacteria in the environment, the status of antimicrobial resistance of Escherichia coli in a variety of wild mammals and their prevalence were examined using antimicrobial-containing media. In total, 750 isolates were obtained from 274/366 (74.9%) wild mammals, and antimicrobial-resistant E. coli was detected in 37/750 isolates (4.9%) from 7 animal species (26/366 [7.1%] individuals). Using antimicrobial-containing media, 14 cefotaxime (CTX)- and 35 nalidixic acid-resistant isolates were obtained from 5 (1.4%) and 17 (4.6%) individuals, respectively. CTX-resistant isolates carried blaCTX-M-27, blaCTX-M-55, blaCTX-M-1, and blaCMY-2, with multiple resistance genes. Fluoroquinolone-resistant isolates had multiple mutations in the quinolone-resistance determining regions of gyrA and parC or qnrB19. Most resistant isolates exhibited resistance to multiple antimicrobials. The prevalence of antimicrobial-resistant bacteria observed in wild mammals was low; however, it is essential to elucidate the causative factors related to the low prevalence and transmission route of antimicrobial-resistant bacteria/resistance genes released from human activities to wild animals and prevent an increase in their frequency.

Burden of Fluoroquinolone Resistance in Clinical Isolates of Escherichia coli at the Ho Teaching Hospital, Ghana.

The growing burden of antibiotic resistance is a threat to the management of infections. Infections by Escherichia coli are routinely treated with fluoroquinolone antimicrobial agents. Due to their frequent use, there has been increasing resistance to these drugs. We set out to determine the burden of fluoroquinolone resistance among clinical E. coli isolates at the Ho Teaching Hospital, Ghana. This was a cross-sectional study conducted from July 2018 to June 2019. One hundred and thirty-five E. coli isolates were cultured from various clinical samples. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disk diffusion method with discs of nalidixic acid (NAL), ciprofloxacin (CIP), norfloxacin (NOR) and levofloxacin (LEV). Deoxyribonucleic acid (DNA) was extracted from the resistant isolates for the detection of fluoroquinolone resistant genes by polymerase chain reaction. Ninety of the 135 isolates (66.7%) were resistant to at least one of the four fluoroquinolone drugs investigated. Resistance to NAL, CIP, NOR, and LEV was 51.0%, 51.1%, 38.8% and 35.7% respectively. Out of the fluoroquinolone resistant isolates, 69 carried one or more fluoroquinolone resistant genes. The predominant resistant genes were aac(6')-Ib-cr (48.9%) and qnrD (25.6%). Seven of the isolates carried both qnrS and aac(6')-Ib-cr genes. Two isolates carried 5 different fluoroquinolone resistant genes. High prevalence of resistance to 4 fluoroquinolone drugs was recorded with associated resistant genes. This is a threat to current efforts to control the spread of antibiotic resistance and calls for concerted efforts to curb the spread of these resistant organisms.