Plasmid-mediated Quinolone Resistance Research Articles

P-1146. Environmental Surfaces in Healthy Households as Reservoirs of Extended-Spectrum Cephalosporin-Resistant Enterobacterales in the Community

Abstract Background Community-acquired extended-spectrum cephalosporin resistant (ESCr)-Enterobacterales (E) are increasing in healthy populations, yet the sources of acquisition are unknown. We assessed household (HH) environmental surface samples (HS) in St. Louis, MO, to identify the surfaces most frequently contaminated with ESCr-E and the associated mechanisms of antibiotic resistance (AR). Figure 1. Enterobacterales within households. Methods From 139 HHs, samples from 21 HS and inguinal fold (IF) samples of healthy adults and children were cultured for bacteria. Families were recruited from outpatient pediatric settings. Bacterial identification and antibiotic susceptibility testing were conducted (Vitek®2, bioMérieux). Molecular characterization of ESCr-E was performed by PCR detection of beta-lactamase (bla) and plasmid-mediated quinolone resistance (PMQR) genes. Whole genome sequencing (WGS) was performed on select HH with > 1 colonized HS to evaluate for genetic relatedness, plasmids, and AR genes. A HH cluster was defined as < 15 single nucleotide polymorphism differences between 2 isolates. Figure 2. ESCR Enterobacterales by genera Results Of 581 IF and 2801 HS, the majority harbored bacteria (IF, 97-100% and HS, 79-100%) and gram-negative bacteria, (IF, 72-80% and HS, 75-87%). IF colonization with Enterobacterales was ∼23% and ranged from 7-60% on HS; The most common ESCr-E genera were Enterobacter (79%), Pantoea (10%), and Klebsiella (7%). ESCr-E were found in 2% of IF. HS most notably colonized with ESCr-E were the kitchen sink faucet (9%), oven door handle (4%), sofa (4%), bathroom sink faucet (3%) and bed sheets (3%), (Figures 1-3). ESBL-E were recovered from ∼2% of HHs. MDR-E (> 3 antibiotic classes) were found in 4% of HHs. WGS of 20 E. cloacae from 4 HHs revealed within HH clusters (Figure 4). Of interest, we found that among E. cloacae, 10/20 (50%) contained AR plasmids (ex. IncFI, IncFII, IncX5, IncHI2, Col440I); bla, other AR (ex. PMQR, aminoglycoside, sulfa, fosfomycin) and virulence genes were detected in E. cloacae in all HHs. We also found related E. cloacae strains in 2 HHs (ST108), indicating local strains circulating potentially related to a common exposure. Figure 3. AR Enterobacterales by surface. Conclusion ESCr-E are present in healthy household environments. Future directions will be to identify candidate clinical and epidemiological features linked with ESCr-E HH colonization. Figure 4. Genetic relatedness among household Enterobacter cloacae Disclosures Ahmed Babiker, MBBS, Beckman Coulter Inc.: Advisor/Consultant

P-1061. In vitro Activity of Gepotidacin against Klebsiella pneumoniae, Including Molecularly Characterized Fluoroquinolone not Susceptible Subsets Causing Urinary Tract Infections in the United States (2019-2022)

Abstract Background Gepotidacin (GEP) is a novel, bactericidal, first-in-class triazaacenaphthylene antibiotic that inhibits bacterial DNA replication by a unique mechanism of action, distinct binding site and provides well-balanced inhibition (for most uUTI uropathogens) of two different type II topoisomerase enzymes. This study reports the activity of GEP and other oral antibiotics against K. pneumoniae, including molecularly characterized fluoroquinolone (FQ) not susceptible (NS) isolates collected from urinary tract infection (UTI) patients in the US. Methods A total of 2,001 K. pneumoniae were collected from 73 sites. CLSI methods were used for both susceptibility (S) testing and MIC interpretation. Isolates with MIC≥0.5 mg/L for ciprofloxacin (NS) and/or ≥1 mg/L for levofloxacin (NS) were screened for FQ-resistance (R) mechanisms (plasmid-mediated quinolone resistance [PMQR]), QRDR mutations, and expression of acrAB and oqxAB by qRT-PCR. Results GEP inhibited 94.9% of all these isolates at ≤16 mg/L (MIC50/90 = 4/16mg/L). GEP MIC50/90 against FQ-S isolates was 4/8 mg/L, and other agents showed activity against ≥90% of isolates, except nitrofurantoin (37.4%S). 14.5% (291/2,001) of isolates were screened for FQ-R mechanisms. GEP MIC50/90 against FQ-NS strains was 16/32 mg/L, whereas other agents had S of 5.8–48.5%. GEP had generally similar activity (MIC50/90, 16-32/32-64 mg/L) against isolates with wildtype QRDR and with or without PMQR genes. Most of the isolates (95.2%) without PMQR genes overexpressed oqxAB and/or acrAB, against which amoxicillin-clavulanate and cefazolin (90.3–95.2%S) were active. GEP had consistent MIC90 of 16 mg/L against isolates with distinct QRDR mutations and absence of PMQR genes, whereas other agents had S of ≤68%. Conclusion GEP showed activity against FQ-S and FQ-NS K. pneumoniae UTI isolates from the US, in particular against FQ-NS isolates carrying QRDR mutations, where standard oral antibiotics showed limited activity. GEP MICs were not substantially affected by QRDR mutations. Highest MICs were seen against FQ-NS isolates with wildtype QRDR and PMQR genes and/or overexpression of efflux-pump genes. These data support the development of GEP for the treatment of uUTI caused by K. pneumoniae. Disclosures Rodrigo E. Mendes, PhD, GSK: Grant/Research Support Renuka Kapoor, PhD, GSK: Employee|GSK: Stocks/Bonds (Public Company) Didem Torumkuney, PhD, GSK: Employee|GSK: Stocks/Bonds (Public Company)

Molecular Characterization of Colistin- and Carbapenem-Resistant Klebsiella pneumoniae: mgrB Mutations and Clonal Diversity in Pediatric Intensive Care Isolates.

Colistin- and carbapenem-resistant Klebsiella pneumoniae (ColR CrKp) cause important health problems in pediatric intensive care units (PICUs) due to its ability to harbor multiple resistance genes and spread of high-risk clones. In this study, molecular epidemiological characteristics, transferable resistance genes, and mgrB alterations of ColR CrKp isolated from PICU were investigated. Isolates were identified by MALDI-TOF MS, and antimicrobial susceptibility tests were performed using disk diffusion method, gradient strip test, and broth microdilution method. Extended spectrum beta-lactamase, AmpC beta-lactamase, carbapenemase, 16S rRNA methyltransferase, plasmid-mediated quinolone resistance, and mcr-1 to -5 genes were investigated by polymerase chain reaction. Sanger sequencing was performed to obtain blaOXA-48-like and mgrB sequences. Pulsed-field gel electrophoresis and multilocus sequence typing were used to determine the clonal spread of the isolates. Ten ColR CrKp harboring blaOXA-48 (70%), blaOXA-232 (20%), blaCTX-M (90%), armA (20%), qnrB (20%), and qnrS (50%) were identified. No mcr genes were found, whereas mgrB mutations through modifications (A7T, C88T, and A121G) and insertion of an IS-1-like insertion sequence were determined. Isolates belonged to ST 14, ST 37, ST 101, ST 147, ST 661, ST 985, and ST 2096. It is crucial to determine the antimicrobial resistance properties and the clonal spread of the isolates to guide the treatment decisions, implement effective infection control measures, and develop novel antimicrobial strategies.

Deciphering spread of quinolone resistance in mariculture ponds: Cross-species and cross-environment transmission of resistome.

Mariculture is known to harbor antibiotic resistance genes (ARGs), which can be released into marine ecosystems via oceanic farming ponds, posing a public health concern. In this study, metagenomic sequencing was used to decipher the profiles of quinolone-resistant microbiomes and the mechanisms of quinolone resistance in sediment, seawater, and fish gill samples from five mariculture ponds. Residues of both veterinary-specific (enrofloxacin and sarafloxacin) and prohibited quinolones (ofloxacin, ciprofloxacin, pefloxacin, norfloxacin, and lomefloxacin) were detected. We identified a total of 285 subtypes of ARGs across all samples. Pathogens played a crucial role in the prevalence and distribution of these ARGs. Out of the annotated 629 bacterial species, 42 were identified as pathogenic, predominantly belonging to the Proteobacteria phylum. Notably, the Acinetobacter genus was prevalent in the gills and exhibited correlations with various ARGs. The presence of the plasmid-mediated quinolone resistance (PMQR) genes in various bacterial species and the identification of sulfonamide resistance genes across different samples indicated the potential for cross-species and cross-environment transmission of ARGs. Metagenomic binning revealed that Exiguobacterium harbored five ARGs (vanA, vanB, fexA, msr(G), mefF), while Shewanella carried six ARGs (blaOXA-436, adeF, qacl, ANT (2'')-Ia, dfrA1, rsmA). Mutations in gyrA and parC contributed to quinolone resistance in these multidrug-resistant Exiguobacterium and Shewanella. Our findings suggest a potential for ARG transmission across various bacterial species and environments in mariculture. This study emphasized the risk of resistance spread within the mariculture ecosystem.

Clonal Spread and Genetic Mechanisms Underpinning Ciprofloxacin Resistance in Salmonella enteritidis.

Salmonella enteritidis is a major cause of foodborne illness worldwide, and the emergence of ciprofloxacin-resistant strains poses a significant threat to food safety and public health. This study aimed to investigate the prevalence, spread, and mechanisms of ciprofloxacin resistance in S. enteritidis isolates from food and patient samples in Shanghai, China. A total of 1625 S. enteritidis isolates were screened, and 34 (2.1%) exhibited resistance to ciprofloxacin. Pulsed-field gel electrophoresis (PFGE) results suggested that clonal spread might have persisted among these 34 isolates in the local area for several years. Multiple plasmid-mediated quinolone resistance (PMQR) genes, GyrA mutations in the quinolone resistance-determining region (QRDR), and overexpression of RND efflux pumps were identified as potential contributors to ciprofloxacin resistance. PMQR genes oqxAB, qnrA, qnrB, and aac(6')-Ib-cr as well as GyrA mutations S83Y, S83R, D87Y, D87G, D87N, and S83Y-D87Y were identified. The co-transfer of the PMQR gene oqxAB with the ESBL gene blaCTX-M-14/55 on an IncHI2 plasmid with a size of ~245 kbp was observed through conjugation, highlighting the role of horizontal gene transfer in the dissemination of antibiotic resistance. Sequencing of the oqxAB-bearing plasmid p12519A revealed a 248,746 bp sequence with a typical IncHI2 backbone. A 53,104 bp multidrug resistance region (MRR) was identified, containing two key antibiotic resistance determinants: IS26-oqxR-oqxAB-IS26 and IS26-ΔISEcp1-blaCTX-M-14-IS903B. The findings of this study indicate that ciprofloxacin-resistant S. Enteritidis poses a significant threat to food safety and public health. The persistence of clonal spread and the horizontal transfer of resistance genes highlight the need for enhanced surveillance and control measures to prevent the further spread of antibiotic resistance.

Quinolone and Colistin Resistance Genes in Extended-Spectrum Beta-Lactamase (ESBL)-Producing Escherichia coli of Diverse Phylogenetic Groups Isolated from Seafood in Mumbai, India

Contamination of coastal-marine environment with multidrug-resistant Escherichia coli has resulted in such bacteria increasingly being detected in the seafood chain. This study aimed to determine the quinolone and colistin resistance genes in extended spectrum-β-lactamase (ESBL)-producing E. coli from seafood. ESBL-producing E. coli isolates (n = 269) were tested for quinolones and colistin resistance phenotypes by disk diffusion and broth microdilution methods, respectively. The isolates were further PCR screened for the plasmid-mediated quinolone resistance (PMQR) genes qnrA, qnrB, and qnrS, genomic mutations in gyrA and parC genes, and the colistin resistance genes mcr-1 and mcr-2. Phylogroup was determined by PCR using the Clermont E. coli phylotyping method. Of 269 isolates tested, 73.60% of E. coli isolates were resistant to moxifloxacin and 8.55% to ofloxacin, the least of all the quinolones tested. Further, 150 (55.76%) E. coli isolates carried at least one of the three PMQR genes tested, where qnrS was the most prevalent gene (53.90%). The colistin resistance gene (mcr-2) was detected in 38 (14.12%) isolates. Twenty-one of these isolates (55.26%) had a colistin minimum inhibitory concentration (MIC) of 16 µg/mL. Based on the Clermont E. coli phylotyping of the isolates harboring at least one of the qnr genes, 66 (44%) belonged to the phylogroup B1, followed by 23 (15.33%) to phylogroup A. Among 38 E. coli isolates carrying colistin resistance gene mcr-2, 27 (71.05%) isolates belonged to phylogroup B1, followed by 4 (10.52%) isolates to phylogroup A. The results suggest that E. coli phylogroups B1 and A harboring plasmid-mediated quinolone and colistin resistance genes are predominant in the seafood supply chain.

Prevalence and antimicrobial resistance of Salmonella isolated from pork in the Northern part of Thailand

Background: Salmonella is one of the most common foodborne bacteria frequently isolated from pork. It pose significant public health risks due to its potential to harbor antimicrobial resistance (AMR) genes. The emergence and spread of multidrug-resistant (MDR) Salmonella strains are of particular concern as these can be transferred to humans through food or contaminated environments. Objective: To determine Salmonella’s prevalence and antimicrobial resistance determinants from pork at retail markets in the Northern part of Thailand. Materials and methods: Salmonella was isolated and identified from 173 pork samples. The isolates were serotyped using the slide agglutination test with somatic- (O) and flagellar- (H) antigens, and tested for antimicrobial susceptibility using 23 different antimicrobials using the disc diffusion method. By polymerase chain reaction (PCR), the extended-spectrum β-lactamases (ESBL) genes (blaTEM, blaSHV, blaCTX-M-1, blaCTX-M-2, blaCTX-M-8/25 and blaCTX-M-9), quinolone-resistance determining region (QRDR) including gyrA, gyrB, parC and parE, plasmid-mediated quinolone resistance (PMQR) determinants including qnrA, qnrB, qnrD, qnrS, and aac(6’)-Ib-cr were determined. Results: In this study, 98.8% of the samples were identified as Salmonella. Among these, 72.5% of isolates showed resistance to at least 1 antimicrobial agent and 48.5% were MDR. Ampicillin/tetracycline/trimethoprim-sulfamethoxazole (AMP/ TE/SXT) was the most common phenotypic resistance pattern. The results revealed that 4 ESBL genes (blaTEM, blaCTX-M-1, blaCTX-M-2, and blaCTX-M-9) were detected in 3 isolates- identified as ESBL producers. Moreover, all 9 quinolone-mediated resistance determinants were observed in quinolone resistance isolates. Conclusion: The results demonstrated that most MDR isolates harbored quinolone resistance determinants. We report the distribution of MDR in Salmonella strains isolated from pork and the correlation between the phenotypic and the genotypic susceptibility. These findings raise great concern about the spread of resistant Salmonella strains in the Northern part of Thailand.

Detection of O25b-ST131 clone in extended spectrum beta-lactamase-producing E. coli from urinary tract infections in Mexico.

Escherichia coli has emerged as an important pathogen in urinary tract infections (UTIs) due to the rapid acquisition of antibiotic resistance genes. This enhances the ability of E. coli to colonize and creates therapeutic challenges within the healthcare system. This study aimed to identify the extended spectrum beta-lactamase (ESBL) and O25b-ST131 pandemic clones in E. coli isolated from two hospitals in Mexico. Bacterial identification and antibiotic susceptibility tests were conducted using the VITEK 2 system. The ESBL and plasmid-mediated quinolone resistance (PMQR) genes were identified by polymerase chain reaction (PCR). E. coli genotyping was carried out by the phylogenetic group analysis and O25b-ST131 identification. A total of 103 unique E. coli clinical isolates were analyzed from a pool of 1,002 strains; 75% obtained from UTIs and vaginal secretions. Multi-resistant antibiotic profiles were observed. Notably, the presence of the aac(6`)lb-cr and qnr genes was associated with 100% ciprofloxacin resistance, when ESBL was present. Additionally, the B2 phylogenetic group was identified, with 23% of isolates belonging to the O25b-ST131 clone. Our research revealed a 10% prevalence of ESBL, in contrast to global prevalence rates. The resistance profiles suggest that the effectiveness of these commonly used antibiotics in treating E. coli-associated UTIs or vaginal infections has decreased significantly. Excessive use of antimicrobial agents contributes to the regional variation. Our results underscore the importance of monitoring the molecular epidemiology, antibiotic resistance, and transmission dynamics of the O25b-ST131 E. coli clone.

Detection of plasmid-mediated quinolone resistance genes in Acinetobacter baumannii clinical isolates in Suez Canal University Hospitals in Ismailia

Detection of plasmid-mediated quinolone resistance genes in Acinetobacter baumannii clinical isolates in Suez Canal University Hospitals in Ismailia

Prevalence of Plasmid-Mediated Quinolone Resistance Genes in Escherichia coli Isolates From Colonic Biopsies of Iranian Patients With Inflammatory Bowel Diseases: A Cross-Sectional Study.

Emerging evidence suggests that ciprofloxacin and other quinolones can be effectively used as adjuncts to immunosuppressive therapy in managing inflammatory bowel disease (IBD). Clinical isolates of Enterobacterales frequently exhibit quinolone resistance. Additionally, increased IBD severity has been linked to the proliferation of Enterobacterales in the gut. This study aimed to explore the frequency of fluoroquinolone resistance and the presence of associated resistance genes in Escherichia coli isolates obtained from intestinal biopsies of patients with IBD in Iran. In this research, we conducted a study that involved the isolation and examination of E. coli bacteria from inflamed ileal and/or colonic tissues of patients diagnosed with IBD, specifically ulcerative colitis (UC) and Crohn's disease (CD), during colonoscopy procedures. We collected demographic and clinical information from the patients. To identify E. coli strains that were resistant to quinolone antibiotics, we performed both phenotypic and molecular analyses. From the colonic and ileal biopsies of 121 patients with IBD, we isolated 107 unique strains of E. coli. Among these strains, 18 (16.8%) were derived from patients with CD, and 89 (83.2%) came from those with UC. Antimicrobial susceptibility tests revealed that 61 out of 107 isolates (57%) of the isolates showed phenotypic resistance to at least one type of quinolone. Additionally, plasmid-mediated quinolone resistance (PMQR) genes, specifically oqxA, oqxB, and qnrS were detected in the E. coli strains linked to both UC and CD. Notably, there was a significant positive correlation observed between intestinal colonization by ciprofloxacin-resistant E. coli and the patients' history of extended ciprofloxacin antibiotic therapy. Our results reveal that a significant number of patients with IBD carry quinolone-resistant E. coli. This colonization may pose a risk factor that could affect disease progression and contribute to potential complications.

Molecular mechanisms impact on fluoroquinolone resistance among E.coli from enteric carriage monitoring before prostate biopsy and earliest description of qnrB81

Fluoroquinolone-resistant (FQs-R) microorganisms causing infectious complications after ultrasound-guided needle biopsy of the prostate (TRUS-BP) have become an important challenge in healthcare settings globally, questioning the continued utility of FQ as the preferred prophylactic agent. This study aimed to characterize molecular mechanisms of resistance on FQs-R E. coli isolated from the enteric microbiota of patients undergoing (TRUS-BP) and to highlight their impact on Minimum Inhibitory Concentrations (MICs). From February 2016 to December 2018, the incidence of rectal carriage of Qs-FQs resistant Enterobacterales detected from rectal swabs of patients before undergoing (TRUS-BP) was 61.06% (80/131) all related to E. coli species. Based on the MICs range of Qs (24–256 mg/L) and FQs (0.24–128 mg/L) breakpoint by EUCAST, we categorized these E. coli isolates into three resistance profiles (I, II, and III) associated with the patterns of chromosomal mutations in the quinolone resistance-determining regions (QRDRs) of gyrA and parC and the plasmid-mediated quinolone resistance encoding genes (PMQRs) detected by PCR-based assay and sequencing; MICs increase in an escalation step according to the co-occurrence of multiple molecular mechanisms. The mutation of the gyrA gene was the most frequent on codons (Ser83Leu/Thr/Tyr/Trp and Asp87Asn); mutation on the parC gene was the least on codons (Ser80Iso/Leu and Glu84 Val/Gly/Lys). PMQRs genes (4 qnrB ,7 qnrS, and one aac(6’)-Ib-cr) were determined within 15% of the isolates. Allelic variation allows us to report earliest the qnrB81 determinant in an E. coli isolate. Among isolates (35%) belonged to the notorious ST131 lineage. The phylogenetic group showed a predominance of B2 group (51, 25%), however (PFGE) revealed a high level of clonal variability. Worrying incidence of FQs-R E. coli isolates in the rectal flora of our local population showed the potential to cause post-infection. FQ resistance is a complex interplay between mutations in the QRDRs and PMQR determinants that impact MICs. The importance of intestinal microbiota as a reservoir of resistant strains and pandemic clones encourages driving mitigation challenges to characterize molecular mechanisms of antimicrobial resistance to adapt prophylactic therapy, control infection, and ensure epidemiological monitoring.

Biofilm Formation and Plasmid-Mediated Quinolone Resistance Genes at Varying Quinolone Inhibitory Concentrations in Quinolone-Resistant Bacteria Superinfecting COVID-19 Inpatients.

The likelihood of antimicrobial failure in COVID-19 patients with bacterial superinfection arises from both phenotypic (biofilms) and genotypic mechanisms. This cross-sectional study aimed to determine the inhibitory concentrations of quinolones-nalidixic acid, norfloxacin, ciprofloxacin, ofloxacin, and levofloxacin-in biofilm formers (minimum biofilm inhibitory concentration [MBIC]) and nonformers (minimum inhibitory concentration [MIC]) and correlate inhibitory concentrations with plasmid-mediated quinolone resistance (PMQR) genes in quinolone-resistant bacteria isolated from COVID-19 inpatients. Quinolone-resistant bacteria (n = 193), verified through disc diffusion, were tested for quinolone inhibitory concentrations using broth microdilution and biofilm formation using microtiter plate methods. The polymerase chain reaction was used to detect PMQR genes. Study variables were analyzed using SPSS v.17.0, with a significance level set at P <0.05. MIC-to-MBIC median fold increases for ciprofloxacin, ofloxacin, and levofloxacin were 128 (2-8,192), 64 (4-1,024), and 32 (4-512) in gram-positive cocci (GPC, n = 43), respectively, whereas they were 32 (4-8,192), 32 (4-2,048), and 16 (2-1,024) in fermentative gram-negative bacilli (F-GNB, n = 126) and 16 (4-4,096), 64 (2-64), and 16 (8-512) in nonfermentative gram-negative bacilli (NF-GNB, n = 24). In biofilm-forming F-GNB and NF-GNB, qnrB (10/32 versus 3/10), aac(6')-Ib-cr (10/32 versus 4/10), and qnrS (9/32 versus 0/10) genes were detected. A 32-fold median increase in the MIC-to-MBIC of ciprofloxacin was significantly (P <0.05) associated with qnrA in F-GNB and qnrS in NF-GNB. Biofilms formed by F-GNB and NF-GNB were significantly associated with the aac(6')-Ib-cr and qnrS genes, respectively. Nearly one-third of the superinfecting bacteria in COVID-19 patients formed biofilms and had at least one PMQR gene, thus increasing the need for quinolones at higher inhibitory concentrations.

Combining with matrine restores ciprofloxacin efficacy against qnrS producing E. coli in vitro and in vivo

The exposure risk of plasmid-mediated quinolone resistance (PMQR) genes increases the incidence of resistant bacterial infections, has resulted in clinical treatment failures with ciprofloxacin, necessitating urgent implementation of novel strategies for controlling this situation. Matrine serves as the principal constituent of the traditional Chinese herb Sophora flavescens Ait. and exhibits pharmacological activities including anti-inflammatory, antibacterial, anti-tumor, and hepatoprotective effects. However, the precise mechanism by which matrine exhibits antibacterial activity remains incompletely elucidated. This study investigated the antibacterial potential and synergistic mechanism of matrine in combination with ciprofloxacin against qnrS-carrying E. coli. The clinical ciprofloxacin-resistant E. coli carrying the qnrS and the recombinant E. coli DE3 (pET28a-qnrS) were evaluated for their antibacterial activity in vitro, it was found that the combination of matrine/ciprofloxacin exhibited a significant synergistic, reducing the MIC value of ciprofloxacin against qnrS-positive E. coli by 4-fold, and it effectively reduced the bacterial load to undetectable levels within 10 h without obvious cytotoxicity. Moreover, consistent findings were observed in significantly reducing bacterial load within the mouse infection model. Molecular docking revealed that matrine was localized in the large loop B of the qnrS crystal structure, establishing hydrogen bonds with Thr-102 and Arg-101, thereby disrupting the activity of qnrS. Interaction analysis further confirmed that matrine could significantly inhibit the protective effect of qnrS on gyrase and restore the activity of ciprofloxacin against qnrS-positive E. coli. Matrine may serve as a qnrS inhibitor to restore the efficacy of ciprofloxacin, suggesting its potential as a novel antibiotic adjuvant for controlling bacterial infections.

Occurrence of Plasmid-Mediated Quinolone Resistance and Carbapenemase-Encoding Genes in Pseudomonas aeruginosa Isolates from Nosocomial Patients in Aguascalientes, Mexico.

Pseudomonas aeruginosa is a leading cause of healthcare-associated infections, which are related to substantial morbidity and mortality. The incidence of Plasmid-Mediated Quinolone Resistance (PMQR) determinants has been previously reported in this bacterium. However, there is limited information regarding the presence of PMQR and carbapenemase-encoding genes simultaneously. This study aims to analyze the prevalence of these determinants on P. aeruginosa strain isolated from clinical patients in the State of Aguascalientes, Mexico. Fifty-two P. aeruginosa isolates from nosocomial patients were collected from Centenario Hospital Miguel Hidalgo. This is a retrospective observational study conducted at a single center. Antibiotic susceptibility was tested using the Vitek-2 system. Only carbapenem-resistant isolates were included in this study. Carbapenemase-encoding genes and PMQR determinants were screened by polymerase chain reaction (PCR). Resistance rates of 100% were found on tigecycline and ceftriaxone. Of the 52 isolates, 34.6% were positive for the qnr genes, 46.2% for the oqxA gene, and 25% for the aac-(6')-lb gene. The most frequent carbapenemase genes found in the samples were blaOXA-51 (42.3%), blaOXA-1 (15.4%), and blaVIM (15.4%). blaOXA-51 co-carrying oqxA was detected in 21.1% of the isolates, blaOXA-51 co-carrying aac-(6')-lb in 11.5%, blaVIM co-carrying aac-(6')-lb in 3.8%, and blaKPC co-carrying oqxA in 5.8%. Systematic surveillance to detect carbapenemase-encoding genes and PMQR determinants, and rational prescription using the last-line drugs could help in preventing the dissemination of multidrug-resistant determinants.

Quinolone and Tetracycline-Resistant Biofilm-Forming Escherichia coli Isolates from Slovak Broiler Chicken Farms and Chicken Meat

Escherichia coli isolates from intensive poultry production are associated with antimicrobial resistance and worldwide health problems. The aim of the study was to detect and evaluate the phenotypic and genotypic antimicrobial resistance, biofilm formation, phylogenetic typing, and virulence factors in E. coli isolates from the rectal swabs of chickens from two farms and swabs of chicken meat purchased from Slovakian food markets. Interpretative readings of minimal inhibitory concentration (MIC) revealed dominant resistance to ampicillin (&gt;50%) in both groups. We also detected higher resistance to ciprofloxacin (45%), tetracycline, ampicillin + sulbactam, and trimethoprim + sulfonamide (each &gt;30%). Here, 28.57% of the strains studied were multidrug-resistant (MDR). The formation of weak biofilms was confirmed in 8.8% of E. coli, while one of the strains obtained from chicken cloacal swabs was classified as a strong biofilm producer. The most frequently confirmed phylogenetic groups in E. coli were B1 and A1 in all groups. PCR detection revealed the presence of genes encoding tetracycline resistance (tetAB) and plasmid-mediated quinolone resistance (qnrABS), and Int1 (52.9%), Tn3 (76.5%), kpsMT II (8.8%), fimA (97.1%), cvaC (38.2%), and iutA (76.5%) genes in the strains studied. Our results demonstrate that chickens and chicken meat were the source of antibiotic-resistant, biofilm-forming, and virulent E. coli, representing a potential risk from the point of view of the One Health concept.

Genomic Characterization of 16S rRNA Methyltransferase-Producing Enterobacterales Reveals the Emergence of Klebsiella pneumoniae ST6260 Harboring rmtF, rmtB, blaNDM-5, blaOXA-232 and blaSFO-1 Genes in a Cancer Hospital in Bulgaria.

Background: Acquired 16S rRNA methyltransferases (16S-RMTases) confer high-level resistance to aminoglycosides and are often associated with β-lactam and quinolone resistance determinants. Methods: Using PCR, whole-genome sequencing and conjugation experiments, we conducted a retrospective genomic surveillance study of 16S-RMTase-producing Enterobacterales, collected between 2006 and 2023, to explore transmission dynamics of methyltransferase and associated antibiotic resistance genes. Results: Among the 10,731 consecutive isolates, 150 (1.4%) from 13 species carried armA (92.7%), rmtB (4.7%), and rmtF + rmtB (2.7%) methyltransferase genes. The coexistence of extended-spectrum β-lactamase (blaCTX-M-3/15, blaSHV-12, blaSFO-1), carbapenemase (blaNDM-1/5, blaVIM-1/4/86, blaOXA-48), acquired AmpC (blaCMY-2/4/99, blaDHA-1, blaAAC-1), and plasmid-mediated quinolone resistance (qnrB, qnrS, aac(6')-Ib-cr) genes within these isolates was also detected. Methyltransferase genes were carried by different plasmids (IncL/M, IncA/C, IncR, IncFIB, and IncFII), suggesting diverse origins and sources of acquisition. armA was co-transferred with blaCTX-M-3/15, blaNDM-1, blaVIM-4/86, blaOXA-48, blaCMY-4, aac(6')-Ib-cr, qnrB, and qnrS, while rmtF1 was co-transferred with blaSFO-1, highlighting the multidrug-resistant nature of these plasmids. Long-read sequencing of ST6260 K. pneumoniae isolates revealed a novel resistance association, with rmtB1 and blaNDM-5 on the chromosome, blaOXA-232 on a conjugative ColKP3 plasmid, and rmtF1 with blaSFO-1 on self-transmissible IncFIB and IncFII plasmids. Conclusions: The genetic plasticity of plasmids carrying methyltransferase genes suggests their potential to acquire additional resistance genes, turning 16S-RMTase-producing Enterobacterales into a persistent public health threat.

Drug-resistant Pantoea agglomerans Causing Bacteremia at a Tertiary Care Hospital in Kolkata, India: First Report of Carbapenem Resistance Mediated by OXA-181.

The spread of antibiotic resistance (ABR) in uncommon human pathogens endangers global public health, escalating morbidity, death, and healthcare expenditures. Pantoea agglomerans, a member of the Erwiniaceae family that rarely infects humans, is emerging as a drug-resistant nosocomial pathogen. Seven P. agglomerans isolates were recovered from bacteremia patients at a tertiary care hospital in Kolkata, West Bengal, between March 2022 and October 2022. The isolates were evaluated for phenotypic resistance, β-lactamase and plasmid-mediated quinolone resistance (PMQR) genes, plasmid profiling, and clonality assessment. All isolates were resistant to fluoroquinolones and third-generation cephalosporins, with four resistant to carbapenems. The following β-lactamases and PMQR genes were identified: blaOXA-1 (n = 1), blaTEM (n = 1), blaCTX-M-1 (n = 2), blaNDM (n = 5), blaOXA-181 (n = 1), qnrB (n = 2), and qnrS (n = 4). Six isolates carried up to seven plasmids ranging in size from 2kb to > 212kb. IncFI, FII, HI, and X3 plasmid types were detected in three isolates, while the rest remained untypable. Four different genetic patterns were noted. Four isolates were clonally related, with three being clonal. The swap of environmental isolates to human pathogens exacerbates the ABR dilemma, periling patient care and outcomes. This is the first report in India of a carbapenem-resistant P. agglomerans blood isolate carrying blaOXA-181. In-depth genomic research of drug-resistant microbes adapted to the environment-human interfaces might underpin the source-route-containment of ABR.

Prevalence of plasmid-mediated quinolone resistance genes in extended-spectrum beta-lactamase producing Klebsiella pneumoniae isolates in northern Iran

Plasmid-mediated quinolone resistance (PMQR) in extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae (K. pneumoniae) contributes to treatment failures, extended hospital stays, and increased mortality percentages. We aimed to determine the prevalence of PMQR genes in ESBL-producing K. pneumoniae isolates from clinical samples in Babol, North of Iran region. This is the first study in this region to investigate this specific association. A total of 95 K. pneumoniae isolates were obtained from hospitalized patients with various clinical infections during March 2022 to February 2023. Disk diffusion and Combination disk method were performed to identification of antimicrobial resistance profiles and ESBL-producing strains. The presence of ESBL and PMQR genes among K. pneumoniae isolates was assessed using polymerase chain reaction (PCR) method. Of the isolates, 68 (71.57 %) were considered as ESBL-producers. The blaTEM, blaSHV and blaCTX-M genes were detected in 74.73 %, 57.89 %, and 41.05 % of K. pneumoniae isolates, respectively. Among the PMQR encoding genes, the highest and lowest frequency was associated to qepA (67.3 %) and qnrA (4.2 %), respectively. The frequency of qnrA, qnrB, qnrS, acc (6′)-Ib-cr, qepA, oqxA, and oqxB genes in 26 MDR-Kp isolates was 11.53 % (n; 3), 69.23 % (n; 18), 65.38 % (n; 17), 73.07 % (n; 19), 80.76 % (n; 21), 84.61 % (n; 22), and 76.92 % (n; 20), respectively. Our result revealed of the 68 ESBL gene-positive isolates, 60 (88.23 %) were positive for the PMQR gene. The co-occurrence of these genes within resistant isolates suggests potential linkage on mobile genetic elements such as plasmids. These findings highlight the significant burden of PMQR determinants in ESBL-producing K. pneumoniae and underscore the urgent need for effective control measures. Implementing robust antimicrobial stewardship programs and strengthening drug-resistance surveillance and control protocols are crucial to prevent the spread of resistant isolates.

Evaluation of extended-spectrum β-lactamase production and plasmid-mediated quinolone resistance in Escherichia coli isolated from raw milk

This study aimed to determine the antibiotic resistance and extended-spectrum β-lactamase production profiles of Escherichia coli isolates obtained from raw milk samples. To this end, the study used a chromogenic-based culture method, MALDI-TOF-MS, 16S rRNA gene sequencing, disc diffusion, and combined disc techniques. PCR was also performed to detect ESBL (blaSHV, blaTEM, and blaCTX-M), quinolone (qnrA, qnrB, and qnrS) genes, and the presence of class I integrons. A total of 47 (78.33%) E. coli isolates were identified by morphological, biochemical, and molecular tests. All isolates exhibited susceptibility to gentamicin and tobramycin. Resistance rates to different antibiotics were observed as follows: ampicillin, 51.06%; nalidixic acid, 23.40%; trimethoprim/sulfamethoxazole, 17.02%; meropenem, 14.89%; ceftazidime, 8.51%; cefotaxime, 8.51%; piperacillin-tazobactam, 4.25%; aztreonam, 4.25%; imipenem, 4.25%; and levofloxacin, 2.12%. ESBL production was positive in 3 (6.38%) of the isolates. Bla genes were detected in 11 isolates (blaSHV = 9 [19.14%], blaTEM = 4 [8.5%], and blaCTX = 3 [6.38%]). The qnrB, qnrS, and qnrA genes were found in 5 (10.63%), 4 (8.51%), and 1 (2.12%) isolates, respectively. The intI1 gene was detected in 14 (29.78%) isolates.

Prevalence of plasmid-mediated quinolone resistance genes and biofilm formation in different species of quinolone-resistant clinical Shigella isolates: a cross-sectional study

BackgroundThe purpose of this study was to look into the presence of plasmid-mediated quinolone resistance (PMQR) genes and biofilm formation in several species of clinical Shigella isolates that were resistant to quinolones.MethodsThe stool samples of 150 patients (younger than 10 years) with diarrhea were collected in this cross-sectional study (November 2020 to December 2021). After cultivation of samples on Hektoen Enteric agar and xylose lysine deoxycholate agar, standard microbiology tests, VITEK 2 system, and polymerase chain reaction (PCR) were utilized to identify Shigella isolates. The broth microdilution method was used to determine antibiotic susceptibility. PMQR genes including qnrA, qnrB, qnrC, qnrD, qnrE, qnrS, qnrVC, qepA, oqxAB, aac(6′)-Ib-cr, and crpP and biofilm formation were investigated in quinolone-resistant isolates by PCR and microtiter plate method, respectively. An enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) technique was used to determine the clonal relatedness of quinolone-resistant isolates.ResultsA total of 95 Shigella isolates including S. sonnei (53, 55.8%), S. flexneri (39, 41.1%), and S. boydii (3, 3.2%) were identified. The highest resistance rates of the isolates were against ampicillin (92.6%, n = 88/95). Overall, 42 of 95 (44.2%) isolates were simultaneously resistant against two or more quinolones including 26 (61.9%) S. sonnei and 16 (38.1%) S. flexneri. All isolates were multidrug-resistant (resistance to more than 3 antibiotics). The occurrence of PMQR genes was as follows: qnrS (52.4%), qnrA and aac(6′)-Ib-cr (33.3%), and qnrB (19.0%). The prevalence in species was as follows: 61.5% and 37.5% (qnrS), 19.2% and 56.3% (qnrA), 38.5% and 25.0 (aac(6′)-Ib-cr), and 19.2% and 18.8% (qnrB) for S. sonnei and S. flexneri, respectively. The other PMQR genes were not detected. In total, 52.8% (28/53) of quinolone-susceptible and 64.3% (27/42) of quinolone-resistant isolates were biofilm producers. Biofilm formation was not significantly different between quinolone-resistant and quinolone-susceptible isolates (P-value = 0.299). Quinolone-resistant isolates showed a high genetic diversity according to the ERIC-PCR.ConclusionIt seems that qnrS, qnrA, and aac(6′)-Ib-cr play a significant role in the quinolone resistance among Shigella isolates in our region. Also the quinolone-resistant S. flexneri and S. sonnei isolates had a high genetic diversity. Hence, antibiotic therapy needs to be routinely revised based on the surveillance findings.