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The rapid rise of antibiotic resistance, combined with the increasing cost and difficulties to develop new antibiotics, calls for treatment strategies that enable more sustainable antibiotic use. The development of such strategies, however, is impeded by the lack of suitable experimental approaches that allow testing their effects under realistic epidemiological conditions. Here, we present an approach to compare the effect of alternative multidrug treatment strategies in vitro using a robotic liquid-handling platform. We use this framework to study resistance evolution and spread implementing epidemiological population dynamics for treatment, transmission, and patient admission and discharge, as may be observed in hospitals. We perform massively parallel experimental evolution over up to 40 d and complement this with a computational model to infer the underlying population-dynamical parameters. We find that in our study, combination therapy outperforms monotherapies, as well as cycling and mixing, in minimizing resistance evolution and maximizing uninfecteds, as long as there is no influx of double resistance into the focal treated community.

Antibiotic-resistant isolates of Salmonella enterica were selected on plates containing lethal concentrations of rifampicin, kanamycin, and nalidixic acid. The stability of the resistance phenotype was scored after nonselective growth. Rifampicin-resistant (Rif ʳ) isolates were stable, suggesting that they had arisen by mutation. Mutations in the rpoB gene were detected indeed in Rif ʳ mutants. In contrast, a fraction of kanamycin-resistant (Km ʳ) and nalidixic acid-resistant (Nal ʳ) isolates showed reduced resistance after nonselective growth, suggesting that mechanisms other than mutation had contributed to bacterial survival upon lethal selection. Single-cell analysis revealed heterogeneity in expression of the porin gene ompC , and subpopulation separation provided evidence that reduced ompC expression confers adaptive resistance to kanamycin. In the case of Nal ʳ isolates, mutations in the gyrA gene were present in most nalidixic acid-resistant isolates. However, the efflux pump inhibitor Phe-Arg-β-naphtylamide (PAβN) reduced the level of resistance in Nal ʳ mutants, indicating that active efflux contributes to the overall level of nalidixic acid resistance. Heterogeneous efflux pump activity was detected in single cells and colonies, and a correlation between high efflux and increased resistance to nalidixic acid was found. These observations suggest that fluctuations in the expression and the activity of critical functions of the bacterial cell, alone or combined with mutations, can contribute to adaptive resistance to antibiotics.

Salmonella enterica serovar Paratyphi A (S. Paratyphi A) has emerged as a significant global health concern due to the progressive development of antimicrobial resistance and its broader geographic distribution. In Taiwan, paratyphoid fever was historically rare and predominantly associated with imported cases. Since 2022, however, a marked increase in domestically acquired infections has been observed, prompting investigations into their origin and likely route of introduction. We analyzed surveillance data on 223 patients with paratyphoid fever reported in Taiwan between January 2001 and December 2024. Whole-genome sequencing and antimicrobial susceptibility testing were performed on 88 S. Paratyphi A isolates obtained from both imported and domestically acquired infections from 2007 to 2024. Phylogenetic analysis and genotyping were conducted to assess genetic relatedness and to trace potential sources of introduction by comparing them with global isolates. Although 55.2% of paratyphoid fever infections were imported, domestically acquired infections became predominant after 2022. Most isolates (76.1%) were resistant to nalidixic acid and nonsusceptible to ciprofloxacin due to gyrA mutations at codon 83 (S83F or S83Y). The majority of domestic isolates were classified as ST129 and paratype 2.4 and showed close genetic relatedness to strains from Indonesia. Of the 31 domestic isolates collected between 2022 and 2024, 30 clustered with Indonesian strains, and 28 exhibited nearly identical genomic profiles, which suggested a prolonged outbreak likely linked to a common external source, such as contaminated imported food. The genomic evidence suggests that the recent increase in domestically acquired S. Paratyphi A infections in Taiwan represents a prolonged outbreak rather than a sustained epidemiological shift. These infections were closely related to strains from Indonesia, suggesting a potential epidemiological link between the two countries in the transmission of paratyphoid fever. While 76.1% of isolates were nonsusceptible to ciprofloxacin due to gyrA mutations, susceptibility to traditional first-line agents remained high. The observed decline in case numbers in 2024 may indicate that the outbreak is subsiding. Genomic surveillance played a crucial role in tracing sources of infection and informing targeted public health responses.

Campylobacter jejuni, a common foodborne zoonotic pathogen, causes gastroenteritis worldwide and is increasingly resistant to antibiotics. We aimed to investigate the antimicrobial resistance (AMR) genotypes of C. jejuni isolated from humans, poultry and birds from wild and urban Italian habitats to identify correlations between phenotypic and genotypic AMR in the isolates. Altogether, 644 C. jejuni isolates from humans (51), poultry (526) and wild- and urban-habitat birds (67) were analysed. The resistance phenotypes of the isolates were determined using the microdilution method with EUCAST breakpoints, and AMR-associated genes and single nucleotide polymorphisms were obtained from a publicly available database. Antimicrobial susceptibility testing showed that C. jejuni isolates from poultry and humans were highly resistant to ciprofloxacin (85.55% and 76.47%, respectively), nalidixic acid (75.48% and 74.51%, respectively) and tetracycline (67.87% and 49.02%, respectively). Fewer isolates from the wild- and urban-habitat birds were resistant to tetracycline (19.40%), fluoroquinolones (13.43%), and quinolone and streptomycin (10.45%). We retrieved seven AMR genes (tet (O), cmeA, cmeB, cmeC, cmeR, blaOXA-61 and blaOXA-184) and gyrA-associated point mutations. Two major B-lactam genes called blaOXA-61 and blaOXA-184 were prevalent at 62.93% and 82.08% in the poultry and the other bird groups, respectively. Strong correlations between genotypic and phenotypic resistance were found for fluoroquinolones and tetracycline. Compared with the farmed chickens, the incidence of AMR in the C. jejuni isolates from the other bird groups was low, confirming that the food-production birds are much more exposed to antimicrobials. The improper and overuse of antibiotics in the human population and in animal husbandry has resulted in an increase in antibiotic-resistant infections, particularly fluoroquinolone resistant ones. Better understanding of the AMR mechanisms in C. jejuni is necessary to develop new strategies for improving AMR programs and provide the most appropriate therapies to human and veterinary populations.

Salmonella Heidelberg is commonly reported in foodborne outbreaks around the world, and chickens and poultry products are known as important source of these pathogen. Multidrug-resistant S. Heidelberg strains are disseminated into poultry production chair, which can lead to severe clinical infections in humans and of difficult to treat. This study aimed at evaluating the β-lactam susceptibility and genotypic relatedness of Salmonella Heidelberg at Brazilian poultry production chain. Sixty-two S. Heidelberg strains from poultry production chain (poultry, poultry meat and poultry farm) were used. All strains were evaluated to antimicrobial susceptibility by diffusion disk test, as well as β-lactam resistance genes. Genotypic relatedness was assessed by Pulsed-Field Gel Eletrophoresis, using Xba1 restriction enzyme. Forty-one strains were characterized as multidrug-resistant according to phenotype characterization. The resistance susceptibility revealed 31 distinct profiles, with higher prevalence of streptomycin (61/62), nalidixic acid (50/62), tetracycline (43/62) and β-lactam drugs (37/62). blaCMY-2 was the more frequent β-lactamase gene found (38/62); other resistance genes found were blaCTX-M (2/62), blaSHV (3/62) and blaTEM-1 (38/62). No carbapenemase genes was found. The Pulsed-Field Gel Electrophoresis showed 58 different profiles. Strains with a larger number of antimicrobial resistance were grouped into ten major clusters apart from others. The spread of resistance by ampC continues to rise, thereby turning concern to public health, since the β-lactam antimicrobials are used as a therapeutic treatment in humans.

Synergistic combinations of antimicrobial agents with different mechanisms of action have been introduced as more successful strategies to combat infections involving multidrug resistant (MDR) bacteria. In this study, we investigated synergistic antimicrobial activity of Camellia sinensis and Juglans regia which are commonly used plants with different antimicrobial agents. Antimicrobial susceptibility of 350 Gram-positive and Gram-negative strains belonging to 10 different bacterial species, was tested against Camellia sinensis and Juglans regia extracts. Minimum inhibitory concentrations (MICs) were determined by agar dilution and microbroth dilution assays. Plant extracts were tested for synergistic antimicrobial activity with different antimicrobial agents by checkerboard titration, Etest/agar incorporation assays, and time kill kinetics. Extract treated and untreated bacteria were subjected to transmission electron microscopy to see the effect on bacterial cell morphology. Camellia sinensis extract showed higher antibacterial activity against MDR S. Typhi, alone and in combination with nalidixic acid, than to susceptible isolates.\" We further explore anti-staphylococcal activity of Juglans regia that lead to the changes in bacterial cell morphology indicating the cell wall of Gram-positive bacteria as possible target of action. The synergistic combination of Juglans regia and oxacillin reverted oxacillin resistance of methicillin resistant Staphylococcus aureus (MRSA) strains in vitro. This study provides novel information about antimicrobial and synergistic activity of Camellia sinensis and Juglans regia against MDR pathogens.

Objective: To investigate the prevalence of Salmonella Typhi and Paratyphi resistance to quinolones and characterize the underlying mechanism in Jiangsu Province of China. Methods: Antimicrobial susceptibility testing was performed using Kirby-Bauer disc diffusion system. Quinolone resistance-determining region (QRDR), plasmid-mediated quinolone resistance (PMQR) determinant genes were detected by PCR and sequencing. Results: Out of 239 Salmonella isolates, 164 were S . Typhi and 75 were S . Paratyphi. 128 (53.6%) Salmonella isolates were resistant to nalidixic acid; 11 (4.6%) isolates to ciprofloxacin and 66 (27.6%) isolates were intermediate to ciprofloxacin. QRDR were present in 69  S . Typhi isolates, among which mutation at codon 83 ( n  = 45) and 133 ( n  = 61) predominated. In S . Paratyphi, the most common mutations were detected in gyrA at codon 83( n  = 24) and parC : T57S ( n  = 8). Seven mutations were first reported in Salmonella isolates including gyrB : S426G, parC : D79G and parE : [S498T, E543K, V560G, I444S, Y434S]. PMQR genes including qnrD1 , qnrA1 , qnrB4 , aac (6 ′ )-Ib-cr4 and qnrS1 were detected in 1, 2, 3, 7 and 9 isolates, relatively. Conclusions: High resistance to quinolones in Salmonella remains a serious problem in Jiangsu, China. The presence of the novel mutations increases the complexity of quinolone-resistant genotypes and poses a threat to public health. Subject terms: Salmonella Typhi, Salmonella Paratyphi, antimicrobial resistance, QRDR, PMQR.

The presence of antibiotic pollutants in water and wastewater can cause significant risks to the environment in different aspects. Therefore, antibiotics need to be removed from water. This study investigates the adsorption of nalidixic acid (NA), a common antibiotic, using bovine serum albumin nanoparticles (BSA NPs). These NPs were synthesized via desolvation technique and characterized using SEM, DLS, FT-IR, and UV-Vis spectroscopy. The effects of adsorbent dosage (0.02–0.9 mg), initial NA concentration (30–80 mg L − 1 ) and contact time (0.5–24 h) on adsorption efficiency were considered. Adsorption isotherms and kinetics were determined experimentally. The Freundlich isotherm best described the adsorption equilibrium, while the pseudo-second-order kinetic model accurately represented the adsorption process. Thermodynamic parameters confirmed the spontaneous and exothermic nature of NA adsorption onto BSA NPs. Under optimal conditions, BSA NPs achieved a removal efficiency of 75% for NA with a maximum adsorption capacity of 240 mg g − 1 . These results demonstrate the potential of BSA NPs as an effective adsorbent for removing NA from aqueous solutions.

Shigellosis is the second leading cause of diarrheal death globally. The global burden has been complicated by the emergence of Shigella strains resistant to first line antibiotic treatments such as ciprofloxacin. This study aims to describe the epidemiologic distribution of the most common Shigella species, and their antimicrobial susceptibility patterns to ciprofloxacin and nalidixic acid (NA) in Latin America. Laboratory data from 19 countries were obtained through the Latin American Network for Antimicrobial Resistance Surveillance (ReLAVRA) from 2000-2015. The Clinical Laboratory Standards Institute reduced susceptibility breakpoints for Enterobacteriaceae was used to interpret the disc diffusion tests for Shigella susceptibility to ciprofloxacin and NA. Negative binominal regression was used to analyze longitudinal trends of Shigella isolates antimicrobial susceptibility. 79,548 Shigella isolates were tested and reported between 2000-2015. The most common isolated species were S. flexneri (49%), and S. sonnei (28%). There was a steady increase in the proportion of S. sonnei isolates within the region(p<0.001). The average annual percentage increase (AAPI) in nonsusceptibility was 18.4% (p<0.001) for ciprofloxacin (baseline = 0.3); and 13.2%(p<0.001) for NA (baseline = 3). AAPI nonsusceptibility to ciprofloxacin was 13.3% for S. flexneri (p<0.04); and 39.9% for S. sonnei (p<0.001). Honduras, Dominican Republic, Venezuela, and Chile reported the highest increase in nonsusceptibility to ciprofloxacin among all Shigella isolates. There is an increasing trend in Shigella nonsusceptibility to ciprofloxacin and NA, including among the most common shigella species, in Latin America. This rise of nonsusceptibility among Shigella species to commonly used treatments such as ciprofloxacin is alarming and threatens the control and management of this currently treatable infection. Improved data quality, collection and reporting is needed in Latin America to respond effectively to the rising trends observed. This includes the need for quality isolate level epidemiological data; molecular data, and data on antibiotic consumption and use.

Background Water is considered a source for the transmission of Arcobacter species to both humans and animals. This study was conducted to assess the prevalence, distribution, and pathogenicity of A. butzleri strains, which can potentially pose health risks to humans and animals. Cultures were isolated from surface waters of a mixed-use but predominately agricultural watershed in eastern Ontario, Canada. The detection of antimicrobial resistance (AMR) and virulence-associated genes (VAGs), as well as enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) assays were performed on 913 A. butzleri strains isolated from 11 agricultural sampling sites. Results All strains were resistant to one or more antimicrobial agents, with a high rate of resistance to clindamycin (99%) and chloramphenicol (77%), followed by azithromycin (48%) and nalidixic acid (49%). However, isolates showed a significantly ( p  < 0.05) high rate of susceptibility to tetracycline (1%), gentamycin (2%), ciprofloxacin (4%), and erythromycin (5%). Of the eight VAGs tested, cia B, mvi N, tly A, and pld A were detected at high frequency (> 85%) compared to irg A (25%), hec B (19%), hec A (15%), and cj 1349 (12%) genes. Co-occurrence analysis showed A. butzleri strains resistant to clindamycin, chloramphenicol, nalidixic acid, and azithromycin were positive for cia B, tly A, mvi N and pld A VAGs. ERIC-PCR fingerprint analysis revealed high genetic similarity among strains isolated from three sites, and the genotypes were significantly associated with AMR and VAGs results, which highlight their potential environmental ubiquity and potential as pathogenic. Conclusions The study results show that agricultural activities likely contribute to the contamination of A. butzleri in surface water. The findings underscore the importance of farm management practices in controlling the potential spread of A. butzleri and its associated health risks to humans and animals through contaminated water. Highlights 1. Arcobacter butzleri was highly prevalent in agricultural surface water. 2. Strains were highly resistant to antimicrobial agents. 3. Virulence genes were detected at a high frequency. 4. High genetic similarity and association among strains from various sites.