Rapid detection of fluoroquinolone-resistant Staphylococcus epidermidis and its associated mutations in the quinolone resistance-determining region.

This study aimed to genomically characterize clinical Enterococcus faecalis and Staphylococcus epidermidis isolates obtained from aspirate and sputum samples at Bayburt State Hospital, focusing on antimicrobial resistance genes, virulence factors, and mobile genetic elements. Whole-genome sequencing (WGS) was performed on the isolates, and antimicrobial resistance genes, virulence determinants, multilocus sequence types (MLST), plasmid replicons, and mobile genetic elements were identified using bioinformatics tools. The E. faecalis E5 (ST64) isolate, recovered from an aspirate sample, carried multiple antimicrobial resistance genes, including mecA, vanT, erm(B), tet(M), and qacA, along with a diverse virulence gene repertoire (cyl, ebp, ace, and agg), mobile genetic elements (Tn6009, ISL1a3), and four plasmid replicons (rep19C, rep9C, repUS43, and rep7a). The E. faecalis E184 and E187 isolates (ST124), obtained from sputum samples, harbored fewer acquired resistance genes but shared gyrA (T672A) and parC (V307I) mutations associated with fluoroquinolone resistance. The S. epidermidis ST59 isolate carried resistance genes lnu(A), fosB, and qacD, together with the plasmid replicon rep7a and insertion sequence ISSau4. The co-occurrence of lnu(A) and the rep7a plasmid in both species suggests a potential pathway for interspecies transmission of resistance determinants. This study reports the genomic detection of mecA in an E. faecalis isolate from Türkiye based on WGS analysis. However, this finding is based solely on genomic data and lacks phenotypic or experimental validation. The detection of a shared plasmid-associated resistance element (lnu(A)-rep7a) in both E. faecalis and S. epidermidis highlights the potential for interspecies resistance gene flow within the hospital environment.

Neisseria gonorrhoeae is a major public health concern due to its high global prevalence and rapid evolution of antibiotic resistance. A first-in-class topoisomerase inhibitor, zoliflodacin (a spiropyrimidinetrione) recently received FDA approval for treatment of gonorrhea, but its potential for cross-resistance with another topoisomerase inhibitor, the fluoroquinolone antibiotic ciprofloxacin, remains poorly understood. Here, we investigated how genetic diversity in the fluoroquinolone target gyrA influences the resistance and fitness effects of the zoliflodacin resistance mutation gyrB D429N . We constructed an isogenic panel of N. gonorrhoeae to determine how the resistance and fitness effects of the gyrB D429N mutation are modulated by the most common ciprofloxacin resistance-associated variants in gyrA . In the presence of gyrB D429N , the zoliflodacin minimum inhibitory concentration (MIC) was 2-4-fold higher in strains that also contained ciprofloxacin resistance-associated gyrA alleles, and the gyrB D429N mutation reciprocally increased ciprofloxacin MICs of these strains 3-6-fold. Fitness cost of the gyrB D429N mutation varied from modest to severe across gyrA backgrounds, with the largest cost in ciprofloxacin resistant gyrA 91F/95G and gyrA 91F/95N backgrounds and comparatively minimal cost in the ciprofloxacin resistant gyrA 91F/95A background. These results demonstrate the capacity for epistatic interactions among resistance-associated gyrA and gyrB mutations, underscoring the need for genomic surveillance to monitor high-risk combinations of resistance determinants as new therapies are deployed.

Shigella is a highly infectious diarrheal pathogen in developing countries. The increasing number of MDR isolates is an emerging problem in resource-limited settings such as India. Detecting resistance genes and virulence genes of Shigella species can help plan empirical therapy and aid in future vaccine development. This study was conducted to decipher the virulence and antimicrobial resistance among our Shigella isolates using targeted PCR. The antimicrobial susceptibility testing of all Shigella isolates was done as per the CLSI 2021 using the Kirby-Bauer disc diffusion method. The genomic DNA was subjected to PCR for the detection of resistance genes and virulence genes. The contig sequence of positive amplicons was compared and analysed using BLAST (Basic Local Alignment Search Tool, NCBI) and the translated protein by ExPASy (Expert Protein Analysis System) server. Sequence determination of the QRDRs of gyrA, gyrB, parC and parE genes showed mutations in gyrA and parC. In GyrA at Gln69/Trp, Phe71/Ser, Ser72/Pro, Met75/Leu, Ser90/Cys, Met94/Leu, His106/Pro, Asn161/His, Thr163/Ala and in ParC at Ala64/Asp; aac-(6')Ib-cr were detected in 95.9% (215/224) and qnrS in 71.8% (161/224) of the isolates respectively. Bla_OXA was found in 92.8% (208/224), while bla_TEM in 37.9% (85/224) and bla_CTX in 8.0% (18/224); mphA was found in 19.4% (4/21) S.sonnei tested, of which 2 had azithromycin MIC of 64mcg/ml. IpaH was found in all the isolates, but the ompA gene, as well as the outer membrane protein A, was absent in 5 isolates of S.flexneri. The isolates of S.sonnei which showed resistance to azithromycin had IncFII, K and F plasmid types and the mphA gene. The emergence of azithromycin resistance among Shigella is a major challenge in controlling multidrug-resistant Shigella. The absence of the ompA protein in some of our isolates limits its use as a potential vaccine candidate.

Gulls (Larus spp.) are key reservoirs and vectors of antimicrobial resistance (AMR). We characterized extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpCproducing Escherichia coli from Great Black-backed (Larus marinus) and American Herring (Larus smithsonianus) gulls in Nova Scotia, Canada. From 100 fecal samples, 34 cefotaxime-resistant isolates were recovered: 29 ESBL and 5 AmpC producers. Detected, and blaCMY-2 + blaTEM-1C (n = 1).. Additional resistance genes conferred resistance to sulfonamides (sul2/sul3), trimethoprim (dfrA14/dfrA17), tetracyclines (tet(A)/tet(B)), aminoglycosides, chloramphenicol (floR), macrolides (mph(A)), lincosamides (lnu(F)), and quinolones (qnrS), complemented by chromosomal mutations in gyrA, parC, and parE. The isolates represented phylogroups A (n=10), B1 (n=18), and D (n=6), and high-risk sequence types including ST155 (n=12), ST744 (n=7), ST224 (n=4), and ST648 (n=3). Plasmid analysis revealed a high replicon burden (≥3 in 70.6% of isolates), primarily IncFIB (70.6%), Col (58.8%), and IncI (50.0%). Key virulence factors detected were fimH (97.1%), lpfA (73.5%), fyuA (47.1%), and iss (38.2%). These results demonstrate that synanthropic gulls in Atlantic Canada carry multidrug-resistant ESBL/AmpC E. coli from globally significant clones, underscoring their role in the dissemination of antimicrobial resistance within a One Health context.

In the studied region, primary resistance to clarithromycin and quinolones in Helicobacter pylori exceeds 65% and rises sharply with age. Non-invasive fecal PCR or Sanger genotyping of 23S rRNA and gyrA mutations showed ≥85% agreement with invasive E-test results, enabling age-stratified, resistance-guided eradication therapy without endoscopy or culture.

Salmonella enterica serovar Kentucky ST198 is a globally distributed, multidrug-resistant clone with growing concern due to its zoonotic potential. Despite its detection in Bangladesh from poultry, migratory birds, and fish, genomic data from poultry remain limited which is its main reservoir. This study aimed to characterize poultry-derived S. Kentucky ST198 isolates from live bird markets in Dhaka and compare them with strains from migratory birds and fish to assess clonal diversity, resistance profiles, and transmission characteristics. Five isolates recovered from poultry carcasses and slaughterhouse environments from three markets were confirmed as S. Kentucky ST198 by whole genome sequencing. All showed resistance to ciprofloxacin, aminoglycosides, sulfonamides, and tetracyclines, supported by the presence of acquired resistance genes (aadA7, aac(3)-Id, aac(6′)-Iaa, sul1, tet(A), blaTEM-1B) and mutations in gyrA (S83F) and parC (S80I). Core-genome SNP analysis revealed that the five poultry isolates differed by only 0–4 SNPs, indicating recent transmission or a shared source within the poultry distribution network. To explore broader ecological links, these genomes were compared with 15 Bangladeshi ST198 genomes from migratory birds and fish. All genomes showed similar antimicrobial resistance and virulence genes profiles, with variable presence of SPI-4, plasmid replicons and integrons. Combined results from pan-genome analysis, core-genome phylogeny, and principal component analysis (PCA) indicated distinct source-associated clustering: poultry isolates formed a tightly clustered, highly clonal group; migratory bird isolates were more dispersed; and fish isolates formed a separate cluster. The open nature of the pan-genome further suggested ongoing gene acquisition within this population. Globally, Bangladeshi isolates formed a monophyletic clade, indicating a locally maintained lineage circulating across environmental and animal reservoirs. Collectively, these findings represent the local circulation of a clonal MDR S. Kentucky ST198 lineage across poultry and non-poultry sources and highlight the need for genomic surveillance in high-risk settings such as live bird markets.

Non-O1/non-O139 Vibrio cholerae strains are widely distributed in aquatic environments worldwide and are increasingly recognized as potential reservoirs of antimicrobial resistance and virulence-associated determinants. In this study, we performed an integrated phenotypic and genomic analysis of 116 V. cholerae isolates collected in 2024 from environmental and clinical sources in Jiaxing, China, including 106 non-O1/non-O139 isolates, 9 O1 isolates, and 1 O139 isolate. Antimicrobial susceptibility testing showed that most isolates remained susceptible to β-lactam/β-lactamase inhibitor combinations, third-generation cephalosporins, carbapenems, and tigecycline, whereas resistance was more frequently observed for ampicillin, streptomycin, nalidixic acid, and ciprofloxacin. Based on the non-susceptibility criteria of Maitrakas et al., 19 of 116 isolates (16.4%) were classified as multidrug-resistant, whereas none met the definition of extensively drug-resistant. Genomic analysis identified diverse resistance determinants, including plasmid-mediated quinolone resistance genes (qnrVC variants) and quinolone resistance-determining region mutations in gyrA and parC. Virulence-associated genes showed heterogeneous distributions: core regulatory and hemolysis-related genes were highly prevalent, whereas classical cholera toxin genes were largely absent. Several accessory virulence factors, including the RTX toxin operon, chxA, ninth, and makA, were detected in subsets of isolates. Core genome multilocus sequence typing revealed substantial genetic diversity, with environmental and clinical isolates distributed across multiple lineages and showing no clear clustering by isolation source. Overall, these data demonstrate the diverse antimicrobial resistance, virulence-associated gene repertoires, and population structure of the Jiaxing V. cholerae collection, with particular relevance to the predominant non-O1/non-O139 population.

To assess the feasibility, effectiveness, and safety of susceptibility-guided individualised H. pylori therapy in a region without prior resistance data, using a single-arm prospective observational design. This prospective study included patients with confirmed H. pylori infection undergoing gastroscopy at Beijing Huairou Hospital (April 2023-May 2024). Molecular testing targeted 23S rRNA mutations (A2142G, A2142C, A2143G) for clarithromycin (CLA) and gyrA mutations (Asn87→Lys/Thr) for levofloxacin (LVX). Patients were assigned to three susceptibility-guided groups: CLA-sensitive, CLA-resistant+LVX-sensitive, and CLA-resistant+LVX-resistant, each receiving 14-day bismuth-containing quadruple therapy. Eradication was confirmed by ¹³C urea breath test 4-8 weeks post-treatment. Intention-to-treat (ITT) eradication rate was 82.3% (per-protocol [PP]: 87.2%). ITT rates by group: CLA-S 81.8%, CLA-R-LVX-S 83.3%, CLA-R-LVX-R 85.0% (P=0.921). PP rates: 86.3%, 92.1%, 85.0% (P=0.602). No significant differences in compliance or safety (P=0.849; P=0.467). Susceptibility-guided therapy achieved 85-90% eradication regardless of resistance profile. Although the ITT rate (82.3%) fell short of the 90% guideline threshold, it remains clinically acceptable in a moderate-to-high resistance setting (Huairou: CLA resistance 26.7%, LVX 28.8%, dual 8.6%), supporting the value of susceptibility testing.

Campylobacter Contamination and Antimicrobial Resistance in Retail Meat from Vietnam and Japan.

To comprehensively map the phenotypic prevalence, temporal trends, and molecular mechanisms of antimicrobial resistance in Helicobacter pylori isolates among patients in Bangladesh. Scoping review conducted in accordance with the JBI methodology and reported using the PRISMA-ScR checklist. A systematic search was conducted in PubMed, MEDLINE, Scopus, and BanglaJOL to identify peer-reviewed observational studies published between 1 January 2000 and 30 April 2025. Ten studies were reviewed. Metronidazole resistance remained persistently high (77.5%-96.4%), while clarithromycin rates surged from ~10% to > 39%. Levofloxacin resistance also reached critical levels (up to 66.1%). In contrast, amoxicillin (< 10%) and tetracycline (< 20%) retained efficacy. Genomic analysis linked resistance primarily to 23S rRNA (A2142G, A2143G) and gyrA mutations within hpAsia2 and hpEurope lineages. The available evidence indicates a high and increasing burden of H. pylori antibiotic resistance in Bangladesh, particularly to clarithromycin, metronidazole, and levofloxacin. These findings highlight the need for updated, evidence-informed treatment strategies and expanded molecular surveillance to support effective eradication therapy.

Rifampicin-resistant Mycobacterium tuberculosis (MTB), a critical priority pathogen, becomes highly complicated when it develops into pre-extensively drug-resistant (pre-XDR) and is an exceptional challenge for global TB control efforts. This study aims to determine the frequency and mutations associated with fluoroquinolone (FLQ) and aminoglycoside (AMG) resistance in MTB isolates from patients with pulmonary illnesses in Kaduna State, Nigeria. A total of 144 MDR-TB-positive sputum samples previously collected from 360 individuals were processed using the NaOH-Na-citrate-NALC method. All isolates were tested for MTB complex using TB Ag MPT64 (SD Bioline), and GenoType MTBDRsl VER 2.0 was used to identify chromosomal mutations in gyrA, gyrB, rrs and eis genes. Risk associated with FLQ- and AMG-resistant MDR-TB was assessed using a structured questionnaire, and the resulting data were statistically analysed. The occurrence rate of FLQ resistance (preXDR) in MDR-TB was 12.5% (n = 18), which was mediated by diverse mutations and missing regions WT1 and WT2 in gyrA (n = 14) and N538D in gyrB (n = 4). The frequency of AMG resistance among MDR-TB strains was 11.8% (7/144), and the predominant mechanisms of resistance were MUT1 (A1401G) and MUT2 (G1484T) in the rrs gene. Moreover, the MUT1 (C-14T) mutation (23.5%, 4/17) was observed in the eis promoter region. Remarkably, among these strains, 2 MDR-TB (1.4%) exhibited resistance to both FLQ and AMG. Histories of contact with TB patients (OR = 8.94, 95% CI: 4.1-19.49, p < 0.0001) and prior anti-tuberculosis treatment (OR = 2.36, 95% CI: 1.13–4.94, p = 0.00233) were associated with FLQ- and AMG-resistant MDR-TB. This study revealed a high occurrence of pre-XDR-TB in the study population and among the MDR strains, which could lead to treatment failures and a higher public health threat. To stop pre-XDR-TB from spreading and growing and to improve treatment outcomes in this and other regions where it is more prevalent, it is imperative to diagnose resistance to second-line anti-tuberculosis drugs quickly before beginning treatment and activate pre-XDR-TB surveillance systems. Not applicable.

Background: Antimicrobial-resistant Escherichia coli and Klebsiella pneumoniae represent an increasing challenge in community-acquired pediatric diarrheal infections. Understanding the genomic basis and dissemination of resistance in outpatient settings is essential for guiding antimicrobial use. Methods: Eighteen Gram-negative isolates obtained from pediatric outpatients with acute diarrhea were analyzed using selective culture methods, antimicrobial susceptibility testing, and whole-genome sequencing. Multilocus sequence typing, serotyping, virulence profiling, antimicrobial resistance gene detection, plasmid replicon typing, mobile genetic element analysis, and core genome-based phylogenetic analysis were performed. Phenotypic resistance profiles were correlated with genomic resistance determinants. Results:Klebsiella pneumoniae (55.56%) and Escherichia coli (44.44%) were identified, with all isolates exhibiting putative multidrug resistance-associated genomic profiles. Extended-spectrum β-lactamase genes, particularly blaCTX-M variants, were strongly associated with resistance to third-generation cephalosporins. In contrast, fluoroquinolone resistance correlated with gyrA and parC mutations and plasmid-mediated qnr genes. Phylogenetic analysis revealed diverse lineages harboring resistance determinants. In silico plasmid analysis revealed that key resistance genes co-occurred with IncF-type plasmids and mobile genetic elements, including ISEcp1, IS26, and class 1 integrons, suggesting putative plasmid association rather than confirmed localization. Conclusions: These findings highlight the small scale of plasmid-mediated antimicrobial resistance among E. coli and K. pneumoniae causing pediatric community-acquired diarrhea. The integration of phenotypic and genomic analyses underscores the need for continuous resistance surveillance to support rational antibiotic use in outpatient settings.

BackgroundThe global emergence of antimicrobial-resistant Neisseria gonorrhoeae represents a major public health concern. In particular, strains carrying the mosaic penA-60.001 allele have been associated with resistance to extended-spectrum cephalosporins and high-level resistance to azithromycin, threatening current treatment strategies. We describe an extensively drug-resistant Neisseria gonorrhoeae (XDR-NG) strain belonging to sequence type (ST) 16406, detected in Northern Italy in April 2025.MethodsThe N. gonorrhoeae strain was recovered from a urethral swab collected from a 28-year-old male at the Microbiology Unit of IRCCS Azienda Ospedaliero-Universitaria di Bologna. Antimicrobial susceptibility testing was performed using E-test strips and interpreted according to EUCAST breakpoints. Whole-genome sequencing (WGS) was conducted using Illumina technology, and genomic characterization included MLST, NG-MAST, NG-STAR typing, resistance gene detection, and phylogenetic analysis with publicly available ST16406 genomes.ResultsThe isolate exhibited high-level resistance to tetracycline (MIC = 16mg/L) and ciprofloxacin (MIC = 4mg/L), very high azithromycin MIC (>256mg/L), and resistance to both injectable and oral ESCs (ceftriaxone MIC = 0.25mg/L, cefixime MIC = 1mg/L). WGS assigned the isolate to MLST ST16406 and identified the mosaic penA-60.001 allele, the A2059G mutation in all four 23S rRNA alleles, mutations in gyrA and parC associated with fluoroquinolone resistance, and tetracycline resistance determinants including tet(M). The A39T mutation was identified in the mtrR repressor gene. No plasmids were detected using PlasmidFinder. Phylogenetic analysis demonstrated close genetic relatedness to previously reported ST16406 isolates from Europe and Asia.ConclusionsThis study reports the first detection in Italy of an XDR N. gonorrhoeae ST16406 isolate carrying the penA-60.001 allele. The finding highlights the ongoing international dissemination of this resistant lineage and underscores the importance of maintaining culture-based diagnostics to ensure effective antimicrobial resistance surveillance.

Antimicrobial resistance and transmission potential of pet-derived Providencia spp. in China.

Wildlife is a critical reservoir of Campylobacter species, particularly C. jejuni and C. coli, carrying diverse genetic lineages, virulence factors, and antimicrobial resistance (AMR) genes. Birds, especially migratory and synanthropic species, are the primary carriers, though mammals, reptiles, and other vertebrates also contribute to maintenance and dissemination. Wildlife-associated strains include both host-specific lineages and generalist clonal complexes (e.g. ST21, ST45, and ST828) capable of crossing wildlife, livestock, humans, and environmental interfaces, reflecting high zoonotic potential. Virulence factors, including motility, adhesion and invasion proteins (CadF, CiaB), and cytolethal distending toxin (CDT), facilitate colonization and survival, while efflux pumps and stress-response genes enhance persistence under antibiotic pressure. AMR is widespread, with resistance to fluoroquinolones, macrolides, tetracyclines, and multidrug phenotypes, driven by anthropogenic contamination, environmental reservoirs, and horizontal gene transfer. Key resistance determinants include gyrA mutations, tet(O), erm(B), cmeABC efflux pumps, and β-lactamases. Despite advances, knowledge gaps remain, particularly for non-avian hosts, environmental reservoirs, and resistance mechanisms. A One Health approach integrating microbiology, genomics, ecology, and epidemiology is essential to map transmission pathways, monitor emerging resistance, and guide interventions to reduce the public health impact of zoonotic and antibiotic-resistant Campylobacter.

Rapid and accurate detection of antimicrobial resistance in Neisseria gonorrhoeae is key for appropriate treatment of gonorrhoea. Nucleic acid amplification tests are increasingly available for the detection of N. gonorrhoeae and antimicrobial resistance markers from clinical samples. This study evaluated the Allplex NG & DR assay for the detection of N. gonorrhoeae and resistance markers against ciprofloxacin (gyrA S91F) and azithromycin (23S rRNA A2059C and C2611T). This was a retrospective diagnostic accuracy study performed in two centres. We evaluated assay inclusivity with a panel of laboratory strains, specificity with 35 N. gonorrhoeae negative clinical samples and sensitivity with 82 N. gonorrhoeae positive samples with a matched culture isolate to allow comparison of resistance calls. The Alinity m STI assay was considered gold standard for N. gonorrhoeae detection, and agar incorporation minimum inhibitory concentration testing of N. gonorrhoeae isolates was considered gold standard for resistance calling. Comparisons were also made to the ResistancePlus GC assay which detects N. gonorrhoeae and the S91F ciprofloxacin resistance marker. The Allplex NG & DR assay performed well in the inclusivity panel and had high sensitivity and specificity for the detection of N. gonorrhoeae in clinical samples (98.8% and 100%, respectively). For resistance detection, sensitivity and specificity were >97%. The Allplex NG & DR assay generated one ciprofloxacin resistance marker false negative due to the presence of the S91I gyrA mutation. Two azithromycin resistance marker false positive calls were identified due to the presence of the 23S rRNA C2611T mutation in phenotypically sensitive cases. Our retrospective study shows the Allplex NG & DR assay performed well in the detection of N. gonorrhoeae and associated azithromycin and ciprofloxacin resistance mutations.

Antimicrobial susceptibility analysis of diarrhoeagenic Escherichia coli isolated from outpatients in Beijing, from 2021 to 2024.

Emergence of a novel cefotaxime- and ciprofloxacin-resistant strain of serogroup Y Neisseria meningitidis sequence type 23 clonal complex in China.

Klebsiella aerogenes is an opportunistic pathogen and a growing cause of healthcare-associated infections, characterized by multidrug resistance and the emergence of global high-risk clones. However, regional genomic surveillance data remain limited. Here, we sought to characterize the population structure, transmission dynamics and resistance mechanisms of clinical K. aerogenes in Albuquerque, New Mexico. We sequenced 177 clinical isolates collected between 2021 and 2023. We also developed a novel, species-specific PopPUNK database to facilitate rapid, high-resolution typing. The New Mexico K. aerogenes population was diverse but dominated by two global pandemic lineages, ST93 (47.5%) and ST4 (7.9%), which were significantly enriched for the virulence factors yersiniabactin and colibactin. Genomic evidence for recent local transmission was rare, with only four putative transmission pairs identified. The resistome was characterized by intrinsic and adaptive mutations. Nearly all isolates possessed gyrA mutations associated with decreased fluoroquinolone susceptibility. Mutations in the AmpC regulator AmpD and the outer membrane porin Omp36 were common, particularly within the dominant ST93 lineage. These mutations have been associated with increased AmpC-mediated carbapenem resistance. Our findings underscore the critical importance of genomic surveillance to monitor the transmission and evolution of adaptive resistance.