Search Results (285)

The incidence of levofloxacin and metronidazole resistance in H. pylori Indonesian strains is increasing. Conventional approaches to antibacterial discovery are often a protracted process. This study uses structure-based virtual screening to quickly discover anti-H. pylori. This study employed homology modeling, docking, ADMET prediction, and molecular dynamics simulation to evaluate phytochemicals against resistant H. pylori gyrA, gyrB, and rdxA structures from Indonesian strains. Three-dimensional structures were constructed from the amino acid sequences gyrA, gyrB, and rdxA of levofloxacin- and metronidazole-resistant H. pylori Indonesian strains. The results of redocking at the binding sites gyrA (0.13 Å), gyrB (0.0024 Å), and rdxA (0.5 Å) obtained a valid RMSD. Curcumin exhibited the lowest average binding scores across gyrA (−154.994 kcal/mol), gyrB (−159.2033 kcal/mol), and rdxA (−166.322 kcal/mol) compared to other compounds and standard therapies, including levofloxacin (−109.1553 and −122.5873 kcal/mol) and metronidazole (−85.6096 kcal/mol). Molecular dynamics simulation results revealed that the curcumin–gyrA complex exhibited comparatively more restrained fluctuation than the other complexes throughout the simulation, as indicated by a consistently low total RMSD value (4–7 Å). Curcumin demonstrated the most favorable computational interaction profile among the evaluated compounds. Full article

Acinetobacter is one of the most relevant pathogenic and nosocomial bacteria in human medicine. However, in veterinary medicine, particularly in Spain, there are very few studies on the impact and frequency of infections due to this genus. The main objective of this study was to characterise Acinetobacter isolates recovered at the Complutense Veterinary Teaching Hospital of the Complutense University of Madrid (HCV-UCM), with special emphasis on detecting antimicrobial resistance. A total of 23 isolates obtained from different animal species and samples over a 25-year period were included in the study, based on their identification as Acinetobacter by VITEK-2. Identification was made by using MALDI-TOF, VITEK-2, whole-genome sequencing (WGS) and a chromogenic medium. WGS confirmed that 13 of the 23 isolates belonged to Acinetobacter spp. Antimicrobial susceptibility was interpreted according to CLSI guidelines using the Kirby–Bauer disk diffusion and broth microdilution method. The proportion of clinical isolates identified as Acinetobacter spp. at the HCV-UCM was 0.3%. Of these, 15.4% (2/13 isolates) were classified as multidrug-resistant. Two isolates with the highest MIC for tigecycline carried the tet(X) gene, and two isolates harboured mutations in both gyrA and parC QRDR regions. The results of this study suggest that, in this hospital, antimicrobial resistance among Acinetobacter isolates may not yet be widespread. Full article

Salmonella enterica serovar Choleraesuis (S. Choleraesuis) is a swine-adapted zoonotic pathogen for which multidrug resistance has become an increasing concern. An antimicrobial resistance profile and multidrug resistance pattern in S. Choleraesuis isolates from diseased pig in Taiwan have been reported. This study was designed as a secondary molecular and statistical analysis of 272 isolates recovered from diseased pigs in Taiwan whose minimum inhibitory concentration phenotypes had been reported previously. Using this isolate collection, we evaluated phenotype–genotype concordance and coresistance patterns by combining antimicrobial susceptibility data with targeted polymerase chain reaction screening and quinolone resistance-determining region sequencing. Multidrug resistance was detected in 97.8% of isolates. Among the screened acquired determinants, blaTEM was the most prevalent gene (94.5%), followed by floR (55.9%) and mcr-1 (12.9%); mcr-3 was detected in only one isolate. Significant phenotype–genotype concordance was observed for aphA-1 with kanamycin resistance and mcr-1 with colistin resistance. Sequence analysis of 60 enrofloxacin-resistant isolates revealed that high-level resistance was associated with combined gyrA and parC substitutions, with additional gyrB-region polymorphisms observed in isolates showing the highest minimum inhibitory concentrations. These findings demonstrate frequent coresistance patterns and complex resistance mechanisms in this lineage and support continued genomic surveillance and prudent antimicrobial stewardship in the swine sector. Full article

The worldwide rise in antimicrobial resistance, along with the ongoing prevalence of cancer, underscores the pressing need for novel, safe, and multifunctional therapeutic candidates. Medicinal plants continue to serve as a valuable source of chemically diverse bioactive molecules that modulate multiple biological targets. In this investigation, the preliminary screening of the antibacterial and anticancer activities of an ethanolic extract of Alhagi maurorum (A. maurorum) was comprehensively evaluated using integrated chemical characterization, in vitro bioassays, and in silico approaches. A liquid chromatography–mass spectrometry (LC-MS) analysis demonstrated a rich phytochemical profile including glucosinolates, phenolic acids, gallotannins, fatty acids, alkaloids, carotenoid derivatives, and 2-hexyldecanoic acid-associated constituents. Antibacterial efficacy was assessed by disk diffusion and minimum inhibitory concentration (MIC) testing against Escherichia coli (E. coli) and Bacillus cereus (B. cereus), with the extract producing inhibition zones similar to those observed with streptomycin. Anticancer effects were determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays with MCF-7 breast carcinoma cells and Hs27 normal fibroblasts over 24, 48, and 72 h, revealing a time-dependent, selective decrease in malignant cell viability with relatively limited toxicity towards normal cells. Induction of apoptosis was further verified by propidium iodide (PI) staining. A molecular docking analysis highlighted 2-hexyldecanoic acid as the principal active compound, with a strong binding affinity for critical bacterial targets (GyrA, GyrB, and RpoB). In silico toxicity and ADME (absorption, distribution, metabolism, and excretion) assessments indicated favorable drug-like properties, good gastrointestinal uptake, and acceptable safety profiles. Altogether, these results provide combined experimental and computational support for A. maurorum as a promising source of dual-purpose antibacterial and anticancer agents and lay a mechanistic foundation for subsequent preclinical studies. Full article

Accurate detection and monitoring of antimicrobial resistance (AMR) in Helicobacter pylori mainly rely on phenotypic methods and culture, which can sometimes fail when bacterial load is low or after recent treatment. We investigated whether gastric biopsies classified as H. pylori-negative by standard diagnostic techniques still contain detectable bacterial DNA, including regions linked to AMR, and assessed whether selected DNA fragments can mediate allelic exchange in vitro. Gastric biopsies from 46 dyspeptic patients in the Democratic Republic of the Congo (including 23 phenotypically positive and 23 phenotypically negative individuals) were analyzed using long-read amplicon sequencing of seven resistance-associated loci, selective whole-genome amplification (sWGA) followed by long-read sequencing of H. pylori-enriched reads, and a proof-of-concept natural transformation assay. Phenotypically negative biopsies exhibited significantly lower sequencing depth across multiple loci (including 23S rRNA, gyrA, gyrB, and pbp1A; p = 0.003–0.014), indicating a reduced H. pylori DNA burden. However, AMR-associated mutations linked to various antibiotic classes were found in both groups. sWGA enabled recovery of fragmentary H. pylori sequence data from phenotypically negative samples, including reads that map to resistance- and virulence-associated genes. In vitro, 23S rRNA A2143G amplicons from both phenotypically positive and negative biopsies produced clarithromycin-resistant transformants in strain 26695. These findings indicate that phenotypically negative gastric biopsies might contain low-abundance and fragmentary H. pylori DNA. Although certain DNA fragments can mediate allelic exchange under controlled in vitro conditions, these results do not confirm bacterial viability, active infection, or clinically relevant in vivo resistance transfer. Therefore, they should be interpreted with caution in molecular AMR surveillance and detection contexts. Full article

Background: Drug-resistant tuberculosis (TB) remains a major challenge in high-burden settings, where timely identification of emerging resistance and effective governance responses are critical. While routine molecular diagnostics generate large volumes of resistance-associated mutation data, these outputs are typically used for individual patient management and remain underutilized for population-level surveillance and for the application of clinical governance approaches for improved TB care. Methods: We conducted a retrospective cross-sectional analysis of 1386 molecular diagnostic records for Mycobacterium tuberculosis, collected between March 2021 and December 2024, from 30 health facilities in the King Sabata Dalindyebo (K.S.D.) Local Municipality of Oliver Reginald (O.R.) Tambo District. Resistance-associated mutation proxies were identified for loci associated with isoniazid (katG, inhA), fluoroquinolone (gyrA), and second-line injectable agents (amikacin, kanamycin, and capreomycin) through mutations in the rrs locus. Mutation proxy prevalence was examined overall, by age group, over time, and across facilities. Persistence of resistance detection was assessed using consecutive-month analyses to characterize temporal continuity at the facility level. Results: At least one resistance-associated mutation proxy was detected in 72.7% of the analyzed records. Isoniazid resistance predominated, with katG mutation proxies identified in 52.2% and inhA in 20.2% of cases. Mutation proxies associated with fluoroquinolone and second-line injectable resistance were less frequently observed. Temporal analysis demonstrated variability over the study period, with a general decline in overall mutation proxy prevalence alongside a relative increase in inhA-associated mutations. Substantial heterogeneity in resistance patterns was observed across health facilities, with high-volume sites contributing the greatest absolute burden and selected facilities demonstrating sustained persistence of mutation detection over consecutive months. These findings highlight the magnitude, distribution, and persistence of resistance-associated mutation proxies within routine programmatic data. Conclusions: Routine molecular diagnostic data revealed a substantial and heterogeneous burden of drug-resistant Mycobacterium tuberculosis in K.S.D. Local Municipality, characterized by age-specific patterns, temporal shifts, and sustained facility-level persistence. Beyond descriptive epidemiology, routinely generated mutation proxy data can serve as early-warning indicators of clinical governance stress, signaling emerging pressures on TB care systems when resistance patterns persist or worsen. Interpreting these trends can support more anticipatory clinical governance, strengthen resistance surveillance, and guide prioritized interventions in high-burden, resource-constrained settings. Full article

Background: Helicobacter pylori causes one of the most prevalent chronic bacterial infections worldwide. Data about H. pylori resistance rates to levofloxacin and rifampin (antimicrobials for second-, third- or fourth-line therapy) in Colombia and South America are scarce. The present study aimed to assess levofloxacin and rifampin resistance rates among 61 H. pylori isolates from the western-central region of Colombia and to identify gyrA point mutations associated with levofloxacin resistance. Methods: For this purpose, antimicrobial susceptibility was determined using the E-test method and gyrA mutations in levofloxacin-resistant isolates were identified by PCR amplification followed by DNA sequencing. Results: Resistance rates to levofloxacin and rifampin were 24.6% and 13.1%, respectively. Among the 15 levofloxacin-resistant isolates, 6 (40%) isolates had ≥2 gyrA mutations and 5 of them exhibited high (≥32 mg/L) levofloxacin MICs. In addition, two new mutations (Y90C and V89D) were also detected. Conclusions: Clinically relevant resistance to levofloxacin and rifampin was detected among H. pylori isolates from central-western Colombia. Multiple gyrA mutations were identified in a significant proportion of levofloxacin-resistant isolates and were mainly associated with high MIC values, highlighting the need for regional surveillance to guide eradication therapies. Full article

Antimicrobial resistance in Escherichia coli (E. coli) is a major public health concern globally, driven by increased resistance to commonly used antimicrobial agents such as β-lactams and fluoroquinolones. The main goal of our research is to develop a machine learning framework to predict antimicrobial resistance in E. coli by integrating antimicrobial susceptibility testing data with genomic biomarker analysis. A dataset comprising 17,122 E. coli clinical isolates was obtained from the Bacterial and Viral Bioinformatics Resource Center (BV-BRC). After preprocessing, fivefold cross-validation was used to train and test five machine learning models: Random Forest, XGBoost, Support Vector Machine, Logistic Regression, and k-Nearest Neighbors. The highest-performing model was XGBoost, with 0.86 accuracy and 0.932 ROC-AUC, followed by Random Forest, with 0.82 accuracy and 0.89 ROC-AUC. Phylogenetic analysis revealed that resistant isolates clustered together relative to the reference genome of E. coli K-12 MG1655. Genomic biomarkers such as gyrA, parC, CTX-M-15, OXA-1, and various multidrug efflux pumps were identified by the Comprehensive Antibiotic Resistance Database (CARD) and ResFinder as significant resistance determinants in this study. In conclusion, this study demonstrates that combining antimicrobial susceptibility testing with machine learning and genomic biomarkers is a powerful framework for predicting antimicrobial resistance in E. coli. Full article

Background/Objectives: The Saker Falcon (Falco cherrug) is an endangered raptor species of ecological and conservation relevance. Despite its status, data regarding its microbiota and the prevalence of antimicrobial resistance (AMR) remain scarce, especially in Eastern Europe. This single-facility study aims to investigate the phenotypic and genotypic AMR profiles of Gram-negative bacteria isolated from captive Saker Falcons in Western Romania. Methods: Freshly voided fecal droppings were collected non-invasively from 40 clinically healthy Saker Falcons. Bacterial identification was performed using selective media and the VITEK^® 2 system. Antimicrobial susceptibility testing (AST) was conducted on a representative subset of 12 isolates. Selected resistance-associated genes were screened by conventional PCR. Results: Escherichia coli was the most prevalent 60% (n = 24/40), followed by Hafnia alvei 10% (n = 4/40) and Pseudomonas spp. 10% (n = 4/40). AST revealed phenotypic resistance among Enterobacteriaceae primarily to ampicillin 20% (n = 2/10), tetracycline 20% (n = 2/10), fluoroquinolones and sulfonamides 10% (n = 1/10), while susceptibility to imipenem 90% (n = 9/10) and gentamicin 90% (n = 9/10) remained high. The targeted resistance-associated genes were detected in selected phenotypically resistant isolates. PCR screening detected blaZ and ampC in 62.5% (n = 5/8) of tested isolates, blaOXA-61 in 37.5% (n = 3/8), blaOXA-51 in 25% (n = 2/8), tetK in 37.5% (n = 3/8), and gyrA in 12.5% (n = 1/8). The isolate used as the negative control, pansusceptible in AST, was confirmed negative for all targeted genes. Conclusions: This single-facility study provides baseline data on AMR traits in Gram-negative bacteria associated with Saker Falcons in Western Romania. Given the limited scale and isolate-based design of the study, the findings should be interpreted cautiously, but they support further investigation of wildlife-associated AMR within a One Health context. Full article

C. pseudotuberculosis is a Gram-positive pathogenic bacterium that infects various animals, causing diseases such as caseous lymphadenitis, leading to significant economic losses in the livestock industry and posing zoonotic risks. This study targeted the conserved gyrA gene fragment of this bacterium, designed specific primers, optimized the reaction system and conditions, and established a high-resolution melting curve (HRM) detection method with potential utility for preliminary molecular screening. Validation showed that this method exhibits strong specificity, producing specific amplification only for the target biovars. The melting temperatures (Tm values) for the two biovars were 86.16 ± 0.05 °C and 86.92 ± 0.05 °C, respectively, allowing clear differentiation. It demonstrated high sensitivity, with minimum detection limits of 28 copies/μL and 25 copies/μL for standard plasmids of the ovis and equi biovars, respectively. The method also showed good reproducibility, with intra- and inter-batch coefficients of variation both below 1.0%. Applied to 133 clinical nasal swab samples from goats in Fujian Province, the method detected a positivity rate of 19.5% and indicates that a biovar equi-like gene fragment was detected in goat nasal swabs from Fujian Province via molecular screening. The HRM method developed in this study is sensitive, specific, simple, and cost-effective, enabling rapid detection and biovar differentiation of C. pseudotuberculosis. It is suitable for large-scale clinical sample screening and provides an efficient technical approach for epidemiological monitoring and precise control of the disease. Full article

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. Full article

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. Full article

Pseudomonas aeruginosa is considered a priority pathogen by the World Health Organization due to its resistance to antibiotics. Isolates resistant to ciprofloxacin (CPFX), a bactericide commonly used against P. aeruginosa, usually carry the mutations T83I or D87N in the GyrA subunit of the DNA gyrase. Yet, the molecular mechanisms by which these mutations confer CPFX-resistance to P. aeruginosa are unknown. Here we solved the crystal structure of the P. aeruginosa gyrase catalytic cleavage core and used it to carry out molecular dynamic (MD) simulations of CPFX-gyrase binding in the wild-type as well as the T83I and the D87N mutant systems. Our results show that DNA plays the most prominent stabilizing role once CPFX is bound, with relatively minor contributions from Thr83 or Asp87. Interestingly, we found a solvent cavity adjacent to these residues that may provide CPFX access to the active site. Interaction energy analysis using Umbrella Sampling indicates that Thr83 and Asp87 may influence CPFX trajectory during binding. In the mutant systems, the repulsive potential increases at the cavity site, which may hinder CPFX accessing the binding site. These results shed light on P. aeruginosa resistance to CPFX and may help provide a methodology to identify new therapeutic agents to target fluoroquinolone resistant bacteria. Full article

Tuberculosis (TB) remains a major public health challenge, particularly in sub-Saharan Africa, where the emergence of drug-resistant MTB undermines current control efforts. This study aimed to characterize genetic mutations associated with resistance to first- and second-line injectable drugs in MTB isolates from pulmonary TB patients in the Republic of Congo. A total of 45 culture-positive MTB isolates collected between 2018 and 2019 were analyzed using targeted next-generation sequencing on the MinION device. Resistance-associated genes, including rpoB, katG, embB, gyrA, and rrs, were examined. Overall, 22/45 isolates (48%) harbored at least one mutation associated with drug resistance. Among resistant isolates, rifampicin resistance was detected in 15 of 22 (68.2%), predominantly associated with mutations in the rpoB rifampicin resistance–determining region, including p.Ser531Leu, p.Asp516Val, and p.His526Tyr.Ethambutol resistance was observed in 11 of 22 isolates (50.0%) and was mainly associated with the embB p.Met306Val mutation. Isoniazid resistance was detected in 9 of 22 isolates (40.9%), primarily associated with the KatG p.Ser315Thr mutation. Fluoroquinolone resistance was identified in 2 of 22 isolates (9.1%), mainly associated with gyrA p.Ala90Val and p.Asp94Gly mutations. Resistance to second-line injectable drugs was detected in 1 of 22 isolates (4.5%) and was associated with the rrs p.Ala1401Gly mutation. These findings underscore the need to enhance molecular surveillance and strengthen treatment protocols for drug-resistant TB in the Republic of Congo to effectively combat this evolving health crisis. Full article

There is a growing need to understand ciprofloxacin (CIP) resistance in less prevalent Salmonella serovars like Salmonella Dublin, which causes life-threatening conditions in both humans and animals. This study investigated potential factors contributing to CIP-resistance in a Salmonella Dublin isolate. The isolate was detected from an initial screening of 17 biobanked Salmonella isolates using the Kirby-Bauer disk diffusion (KBDF) method. The minimum inhibitory concentration (MIC) values of the identified CIP-resistant Salmonella Dublin isolate and a CIP-susceptible isolate of the same serovar were also obtained using the broth-dilution (BD) method. The two candidates were then challenged in 1/4 of their respective BD MICs for gene expression analysis, focusing on the acrAB efflux genes and the regulator genes marA, ramA, and soxS. Genomes of the isolates were also sequenced using the Oxford Nanopore sequencing platform, and then analyzed for mutations, antimicrobial resistance genes, and plasmids using ABRicate. The SWISS-MODEL server was used for protein modeling and comparison. For our results, the MIC values (KBDF; BD) for the CIP-resistant and CIP-susceptible Salmonella Dublin isolates were (1.5 μg/mL; 1.95 μg/mL) and (<0.125 μg/mL; 0.03 μg/mL), respectively. Both isolates had genes (mdtK, emrR, emrA, and emrB) notable for fluoroquinolone resistance, with the CIP-susceptible isolate also carrying the IncFII(S) plasmid. Expression of the acrA, acrB, ramA, and soxS genes was markedly higher in the CIP-resistant isolate, which also harbored an Asparagine (N) to Serine (S) mutation at position 868 in the GyrA protein. This mutation, however, caused no significant structural change. Despite reporting on a single CIP-resistant Salmonella Dublin isolate, our result highlights the potentially significant role of an efficient efflux system in contributing to CIP resistance in this isolate, even when no impactful mutations were identified. Full article