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The global tuberculosis (TB) epidemic is becoming progressively more complex due to the increasing prevalence of multidrug-resistant TB (MDR-TB), particularly with resistance to fluoroquinolones (FQs). This study focuses on identifying genetic mutations in the gyrA and gyrB genes of Mycobacterium tuberculosis that drive FQ resistance. Sputum samples from suspected pulmonary TB patients were analyzed using PCR and sequencing to detect mutations within the quinolone resistance-determining regions (QRDR). The analysis revealed that mutations in gyrA, especially S95T, are prevalent and play a key role in FQ resistance. Additionally, less frequent mutations in gyrB, such as E501D and A533P, were also detected. These findings shed light on the molecular mechanisms contributing to FQ resistance in MDR-TB strains and underscore the need for enhanced diagnostic methods to identify resistance patterns more accurately. The insights gained from this research offer a foundation for improving TB treatment approaches and addressing the growing challenge of drug-resistant TB worldwide.

BackgroundThe increasing prevalence of antibiotic resistance in cases of Helicobacter pylori (H. pylori) infection has emerged as a significant global issue. This study offers a comprehensive examination of the alterations in drug resistance exhibited by H. pylori in the Nanjing region of China during the preceding five years. Another important objective is to investigate the influence of levofloxacin medication history on genotypic and phenotypic resistance.MethodsThis research screened 4277 individuals diagnosed with H. pylori infection between April 2018 and May 2023. The phenotype and genotypic resistance were evaluated using the Kirby-Bauer disk diffusion and PCR method.ResultsThe most recent primary resistance rates for metronidazole, clarithromycin, levofloxacin, amoxicillin, furazolidone, and tetracycline were recorded at 77.23% (2385/3088), 37.24% (1150/3088), 27.72% (856/3088), 0.52% (16/3088), 0.19% (6/3088), and 0.06% (2/3088), respectively. For the recent five years, we observed a notable upsurge in the rate of metronidazole resistance and a slight elevation of clarithromycin and levofloxacin resistance. The documented resistance rates to single-drug, dual-drug, triple-drug, and quadruple-drug regimens were 35.39%, 28.32%, 25.72%, and 0.21%, respectively. The prevalence of multidrug-resistant strains escalated, rising from 37.96% in 2018 to 66.22% in 2023. The rate of phenotypic and genotypic resistance rate (57.10% and 65.57%) observed in strains obtained from patients without a levofloxacin treatment history was significantly lower than the rate in strains obtained from those with a history of levofloxacin treatment (88.73% and 94.74%). The prevailing gyrA mutations were primarily N87K (52.35%, 345/659), accompanied by D91N (13.96%, 92/659), and closely followed by D87G (10.77%, 71/659). For gyrA mutations, the 91-amino acid mutants exhibit a higher likelihood of discrepancies between phenotypic and genotypic resistance than the 87-amino acid mutants.ConclusionThe extent of antibiotic resistance within H. pylori remains substantial within the Nanjing region. If levofloxacin proves ineffective in eradicating H. pylori during the initial treatment, its use in subsequent treatments is discouraged. The employment of levofloxacin resistance genotype testing can partially substitute conventional antibiotic sensitivity testing. Notably, predicting phenotypic resistance of levofloxacin through PCR requires more attention to the mutation type of gyrA to improve prediction accuracy.

Xuxia Yu,1 Danyang Zhang,2 Qifa Song3 1Department of Hospital Infection, Ningbo City First Hospital, Ningbo, Zhejiang Province, People’s Republic of China; 2Department of Microbiology, Ningbo Municipal Centre for Disease Control and Prevention, Ningbo, Zhejiang Province, People’s Republic of China; 3Central Laboratory, Ningbo City First Hospital, Ningbo, Zhejiang Province, People’s Republic of ChinaCorrespondence: Qifa Song Email qifasong@126.comPurpose: Fluoroquinolone-resistant Shigella is considered a serious public health problem and has been put on the WHO global priority list of antibiotic-resistant bacteria. This study was aimed to investigate the fluoroquinolone resistance in Shigella and its relevant genetic mechanisms.Materials and Methods: Shigella isolates that were isolated from diarrheal patient’s feces in Ningbo China from 2011 to 2018 were tested for susceptibility to ampicillin, gentamicin, tetracycline, nalidixic acid, ciprofloxacin, and cefotaxime. Genes related to quinolone resistance were amplified by PCR.Results: A total of 118 Shigella isolates were collected, including 76 S. flexneri isolates, 40 S. sonnei isolates, and 2 S. boydii isolates. Ciprofloxacin susceptibility test identified 10 (9%) susceptible, 65 (55%) intermediate, and 43 (36%) resistant isolates. Of 76 S. flexneri isolates, 37 were ciprofloxacin resistant, a prevalence significantly higher than 6 of 40 S. sonnei isolates (P=0.01). The isolates collected during 2014– 2018 displayed a significant increase in the prevalence of ciprofloxacin resistance (P=0.05) than those collected during 2011– 2013. All the ciprofloxacin-intermediate and resistant isolates had mutations of gyrA(S83L) and parC (S80I), whereas only the ciprofloxacin-resistant isolates had gyrA (D87N) mutation and qnrB gene. Additionally, 30% of the ciprofloxacin-resistant isolates were positive for aac(6´)-Ib-cr gene.Conclusion: This study shows the currently increasing prevalence of ciprofloxacin resistance. The reduced fluoroquinolone susceptibility is highly associated with gyrA (S83L) and parC (S80I) mutations, while the fluoroquinolone resistance is highly associated with gyrA (D87N) mutation, qnrB gene and perhaps aac(6´)-Ib-cr gene.Keywords: Shigella, ciprofloxacin resistance, gyrA mutations, plasmid-mediated quinolone resistance, PMQR, quinolone resistance-determining region, QRDR

Noura E Esmaeel, 1 Marian A Gerges, 1 Thoraya A Hosny, 2 Ahmed R Ali, 3 Manar G Gebriel 1 1Medical Microbiology and Immunology Department, Faculty of Medicine, Zagazig University, Zagazig, Egypt; 2Clinical Pathology Department, Faculty of Medicine, Zagazig University, Zagazig, Egypt; 3Urology Department, Faculty of Medicine, Zagazig University, Zagazig, EgyptCorrespondence: Marian A GergesMedical Microbiology and Immunology Department, Faculty of Medicine, Zagazig University, 44519, EgyptTel +20 1003819530Email maromicro2006@yahoo.comIntroduction: Resistance to fluoroquinolones (FQ) in uropathogenic Escherichia coli (UPEC) has emerged as a growing problem. Chromosomal mutations and plasmid-mediated quinolone resistance (PMQR) determinants have been implicated. Data concerning the prevalence of these determinants in UPEC in our hospital are quite limited.Purpose: To investigate the occurrence and genetic determinants of FQ resistance in UPEC isolated from urinary tract infection (UTI) cases in Zagazig University Hospitals.Patients and Methods: Following their isolation, the identification and susceptibility of UPEC isolates were performed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometer (MALDI-TOF MS). FQ resistance was detected by the disc diffusion method. Ciprofloxacin minimal inhibitory concentration (MIC) was determined using E-test. Chromosomal mutations in the gyrA gene were detected using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), and for detection of PMQR, a couple of multiplex PCR reactions were used.Results: Among a total of 192 UPEC isolates, 46.9% (n=90) were FQ resistant. More than half of the isolates (57.8%) exhibited high-level ciprofloxacin resistance (MIC > 32 μg/mL). Mutations in gyrA were detected in 76.7% of isolates, with 34.4% having mutations at more than one site. PMQR determinants were detected in 80.1% of UPEC isolates, with aac(6ʹ)-Ib-cr gene being the most frequent found in 61.1% of isolates.Conclusion: There is a high prevalence of both gyrA mutations and PMQR determinants among UPEC isolates in our hospital which contribute to high-level ciprofloxacin resistance, a finding that may require the revision of the antibiotics used for empirical treatment of UTI.Keywords: gyrA mutations, qnr determinants, uropathogenic E. coli, ciprofloxacin resistance

Abstract Background Nearly 5% of all Mycobacterium tuberculosis strains worldwide are resistant at least to rifampicin and isoniazid (multidrug-resistant tuberculosis, MDR-TB). Inclusion of a fluoroquinolone and an injectable agent (kanamycin, amikacin or capreomycin) in multidrug therapy is crucial for proper treatment of MDR-TB. The incidence of MDR-TB in Kuwait is ~1%. MDR-TB strains additionally resistant to fluoroquinolones and injectable agents are defined as extensively drug-resistant (XDR-TB) strains and have been detected in >55 countries. Infections with XDR-TB strains have very poor prognosis. This study detected the occurrence of gyrA mutations associated with fluoroquinolone resistance among MDR-TB strains in Kuwait. Findings Direct DNA sequencing of quinolone resistance-determining region of gyrA gene was performed to detect fluoroquinolone resistance-associated mutations in 85 MDR-TB strains isolated from 55 TB patients and 25 pansusceptible M. tuberculosis strains. For isolates exhibiting gyrA mutations, 3'-end of rrs (16S rRNA) was sequenced for the detection of XDR-TB. Fingerprinting of fluoroquinolone resistant MDR-TB strains was performed by detecting mutations in three (81 bp hot-spot, N-terminal and cluster II) regions of rpoB, katG codon 315 and inhA-regulatory region, polymorphisms at gyrA codon 95 and katG codon 463 by DNA sequencing and by double-repetitive-element PCR for determining strain relatedness. None of the pansusceptible but six of 85 MDR-TB strains contained gyrA mutations. Only gyrA codon 94 was mutated in all six (D94A in one and D94G in five) strains. Three of six mutant strains were recovered from the same patient while three other strains represented individual patient isolates. Fingerprinting studies identified all individual patient isolates as epidemiologically distinct strains. All six strains with a gyrA mutation contained wild-type rrs sequence. Conclusions Although fluoroquinolones are generally not used for chemotherapy of TB and drug susceptibility testing for second-line drugs is not carried out in Kuwait, four of 55 (7%) individual patient MDR-TB strains contained mutations in gyrA gene. The data advocate routine drug susceptibility testing for this important second-line drug for proper management of MDR-TB in Kuwait. Lack of mutations in 3'-end of rrs gene that confer resistance to injectable agents reduce the likelihood of occurrence of XDR-TB, at present, in Kuwait.