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Topical application of antibiotics in the treatment of orthopaedic implant-related infections can be achieved by using hyaluronic acid (HA)-based hydrogels as carriers. Our aim was to investigate potential toxic effects of a novel antibiotic-loaded hydrogel on osteogenic cells and its antibacterial effect against staphylococci. A covalently cross-linked hyaluronic acid (HA)-based hydrogel was loaded with increasing concentrations of cefuroxime and vancomycin and their release was examined by UV spectrometry. Primary human (HoBs), mouse (MoBs) osteoblasts, or SaoS-2 cells were either exposed to the drug-loaded hydrogel or to antibiotics alone, followed by assessment of cell metabolism and proliferation. Antibacterial effects were evaluated against Staphylococcus aureus (S. aureus) and Staphylococcus epidermidis (S. epidermidis). Increasing concentrations of antibiotics did not affect cell metabolism in any osteogenic cell type, whereas cell proliferation remained unaltered in MoBs, was significantly reduced in SaoS-2, and was stimulated in HoBs. Cultures of MoBs and HoBs tolerated higher concentrations of vancomycin than SaoS-2. Antibiotic-loaded hydrogels did not exert toxic effects on HoBs. After 24 h, 16.8% of vancomycin and 70.8% of cefuroxime were released from the hydrogel. Cefuroxime-loaded hydrogels significantly inhibited growth of S. aureus but not of S. epidermidis, while vancomycin-loaded hydrogel had scarce effects on S. epidermidis. Loading HA-based hydrogel with antibiotics does not harm osteoblasts at clinically relevant concentrations but inhibits bacterial growth. Higher loading of vancomycin may be required due to its slow release while cefuroxime is released more rapidly. A resorbable, antibiotic-loaded hydrogel may be used for implant-related infections in orthopaedics

Introduction. The global spread of extended-spectrum cephalosporinase-producing Escherichia coli (producing extended-spectrum β-lactamase or plasmid-borne AmpC, hereafter ESC-Ec) is a major public health concern. Whilst extensively studied in high-income countries, the transmission pathways between humans and animals in low- and middle-income countries (LMICs) remain unclear. In rural Cambodia, the asymptomatic carriage and transmission dynamics of ESC-Ec between humans and animals living in close proximity are poorly understood, highlighting the need for targeted research in this area.

Gap statement. An enhanced understanding of the genetic epidemiology of ESC-Ec can enable mitigation strategies to reduce the burden of disease and drug-resistant infections in LMIC settings.

Aim. This study aimed to investigate the genetic relatedness and genotypic antibiotic resistance profiles of ESC-Ec strains from humans and livestock in rural Cambodia and to identify patterns of antimicrobial resistance (AMR) gene transmission between hosts and across households and villages.

Methodology. Faecal samples were collected from 307 humans and 285 livestock in 100 households in or near Kampong Cham Province in rural Cambodia. From these samples, 108 ESC-Ec strains were subjected to whole-genome sequencing. Core genome MLST (cgMLST) and phylogenetic analysis determined genetic relationships between strains. All strains were screened for the presence of antibiotic resistance genes and plasmids.

Results. Human and livestock isolates were assigned to six phylogroups, with phylogroup A being the most common (56.5%). MLST identified 50 sequence types (STs), 17 of which were shared between humans and animals, with ST155 being the most prevalent. cgMLST revealed 97 distinct cgMLST sequence types (cgST), indicating strain sharing between humans and animals. Additionally, AMR gene analysis showed widespread resistance, with genes from the blaCTX-M group detected in 84.2% of isolates. Notably, AMR genes such as aph(3'')-Ib–sul2 co-occurred in 50% of isolates. Finally, plasmid analysis identified IncF plasmids in 75.9% of isolates, likely facilitating AMR gene transmission across hosts.

Conclusions. Our findings demonstrate that ESC-Ec strains and their AMR genes are transmitted between humans and livestock in rural Cambodia, likely driven by both clonal spread and plasmid-mediated horizontal gene transfer. These results highlight the urgent need for antimicrobial stewardship and infection control strategies to mitigate the spread of multidrug-resistant pathogens in both human and animal populations.

Urinary tract infections (UTIs), traditionally dominated by Gram-negative pathogens, are increasingly complicated by antimicrobial-resistant Enterococcus spp. in hospital settings, particularly during the use of indwelling catheters. This study screened urine samples from 210 catheterized intensive care unit patients at Uppsala University Hospital (June 2020–September 2021), identifying 39 unique PhenePlate™-RF types across E. faecium, E. faecalis, and E. durans. E. faecium isolates showed considerable diversity, primarily within clonal complex 17 (CC17), known for its virulence and antibiotic resistance. We identified multiple lineages and sequence types (STs), such as in patient HWP143, who had isolates from both ST80 and ST22 (an ancestral CC17 lineage). Notably, metabolic adaptations, such as increased L-arabinose metabolism, and shifts in antibiotic resistance were observed. Variations and similarities in plasmid content between individual lineages suggest horizontal gene transfer. E. faecalis isolates exhibited less diversity, but still significant metabolic variability across patients and mixed infections, as seen in patient HWP051, colonized by both ST16 (CC58) and ST287. E. durans, though less common, shared important metabolic traits with E. faecium and displayed polyclonal characteristics, highlighting its potential role in UTIs and the complexity of enterococcal infections. E. durans was sometimes misidentified, underlining the need for accurate identification methods. This research underscores the importance of understanding genetic and metabolic diversity, plasmid variations, and horizontal gene transfer (HGT) in Enterococcus spp., which influence antibiotic resistance, virulence, and ultimately, treatment outcomes.

Third-generation cephalosporin-resistant E. coli (3GCR-Ec), including extended-spectrum beta-lactamase E. coli (ESBL-Ec), have been detected in environmental water, and are likely discharged by wastewater treatment plants (WWTPs). This study examined the occurrence of 3GCR- and ESBL-Ec in the Swedish urban river Fyrisån, Uppsala, from March to November 2023. Water samples were collected every two months from four sites: upstream of the WWTP (upWWTP), near the WWTP effluent (WWTP), downstream of the WWTP (dWWTP), and Lake Mälaren. Samples were analyzed for 3GCR-Ec and ESBL-Ec using culture- and sequence-based methods. A total of 40 3GCR-Ec isolates were subjected to antibiotic sensitivity testing via microtiter plates and whole-genome sequencing (WGS), followed by in-silico analysis. Significantly higher concentrations of 3GCR-Ec and ESBL-Ec were found at the WWTP compared to Lake Mälaren (P<0.05). When comparing WWTP and upWWTP, significantly higher concentrations of 3GCR-Ec (P<0.05) and borderline significantly higher concentrations of ESBL-Ec (P = 0.0556) were detected at WWTP. All tested E. coli isolates (n = 40) were multidrug-resistant (MDR), with no resistance to imipenem or meropenem. The virulence- and MDR-associated sequence types ST69, ST10, ST1193, and ST1722 were present at multiple sampling points, while ST131 was found only downstream of the WWTP in March and November. Nearly identical strains were detected at WWTP and dWWTP, indicating WWTP as the origin and persistence of resistant strains in the river. Antibiotic-resistance genes (ARGs) were detected in all isolates; 65 % of isolates carried an ESBL gene, and 43 % carried bla_CTX-M-15, common in clinically relevant strains. By tracking resistance dynamics over space and time using WGS, this study highlights the role of WWTPs in the dissemination of antimicrobial resistance in aquatic environments and stresses the need for enhanced surveillance and improved sewage treatment.

Background

Urinary tract infections (UTIs) are prevalent among patients carrying indwelling catheters in the intensive care unit (ICU). This study investigates antibiotic use and bacterial colonisation among ICU patients during the third wave of the COVID-19 pandemic, building on our prior discovery of increased Enterococcus colonisation associated with increased cephalosporin use in early COVID-19.

Methods

Longitudinal urine samples from COVID-19 patients (n = 109) with transurethral catheterisation were analysed for bacterial prevalence, further identified via MALDI-TOF. Microbiological results were combined with clinical data obtained daily, assessed and compared with COVID-19 waves 1 and 2.

Results

Patients in wave 3 exhibited improved outcomes compared to those in waves 1 and 2, alongside a decrease in antibiotic use. Staphylococcus emerged as the primary bacterium and early colonizer of the urinary tract, potentially due to the absence of antibiotic treatment. Our results imply that length of stay (LOS) correlates solely with enteric pathogens and that antibiotic treatment correlates with colonisation by certain uropathogens, whereas the absence of antimicrobial therapy is associated with rapid colonisation of skin flora. Polymicrobial colonisation was common, predominantly involving Gram-positive bacteria.

Conclusion

Our findings underscore the complexity of bacteriuria in ICU patients, advocating for targeted surveillance and tailored antibiotic approaches to mitigate UTI risk. Insights into antibiotic use and bacterial colonisation are vital for optimising stewardship practices, combating antimicrobial resistance, and enhancing ICU patient outcomes.

In vitro screening of large compound libraries with automated high-throughput screening is expensive and time-consuming and requires dedicated infrastructures. Conversely, the selection of DNA-encoded chemical libraries (DECLs) can be rapidly performed with routine equipment available in most laboratories. In this study, we identified novel inhibitors of SARS-CoV-2 main protease (M^pro) through the affinity-based selection of the DELopen library (open access for academics), containing 4.2 billion compounds. The identified inhibitors were peptide-like compounds containing an N-terminal electrophilic group able to form a covalent bond with the nucleophilic Cys145 of M^pro, as confirmed by x-ray crystallography. This DECL selection campaign enabled the discovery of the unoptimized compound SLL11 (IC₅₀ = 30 nM), proving that the rapid exploration of large chemical spaces enabled by DECL technology allows for the direct identification of potent inhibitors avoiding several rounds of iterative medicinal chemistry. As demonstrated further by x-ray crystallography, SLL11 was found to adopt a highly unique U-shaped binding conformation, which allows the N-terminal electrophilic group to loop back to the S1′ subsite while the C-terminal amino acid sits in the S1 subsite. MP1, a close analog of SLL11, showed antiviral activity against SARS-CoV-2 in the low micromolar range when tested in Caco-2 and Calu-3 (EC₅₀ = 2.3 µM) cell lines. As peptide-like compounds can suffer from low cell permeability and metabolic stability, the cyclization of the compounds will be explored in the future to improve their antiviral activity.

The spread of antimicrobial resistance is a global health concern, and resistance mediated by Extended-Spectrum Beta-Lactamases (ESBLs) can cause major consequences. The aim of this study was to explore individuals' perceptions of their daily life and how they cope after being diagnosed with carriage of ESBL-producing bacteria. A qualitative study was conducted with a descriptive design. Data were collected through individual interviews with 24 persons having ESBL carriership, via a semi-structured interview guide. The data were analyzed using qualitative content analysis. The informants' perceptions on "Living with uncertainty about carriership that impacts oneself and others" were interpreted. Experiences of altered behaviors and sentiments due to ESBL carriership were described, as ESBL carriership was perceived to have a psychosocial impact on many informants. Ambiguous and inconsistent information tended to exacerbate these perceptions. The results of this study emphasize the importance of conveying individualized information, both at the time of diagnosis of ESBL carriage and thereafter. This study was not registered.

Bladder urothelial carcinoma (BLCA) is the 10th most common cancer with a low survival rate and strong male bias. We studied the field cancerization in BLCA using multi-sample- and multi-tissue-per-patient protocol for sensitive detection of autosomal post-zygotic chromosomal alterations and loss of chromosome Y (LOY). We analysed 277 samples of histologically normal urothelium, 145 tumors and 63 blood samples from 52 males and 15 females, using the in-house adapted Mosaic Chromosomal Alterations (MoChA) pipeline. This approach allows identification of the early aberrations in urothelium from BLCA patients. Overall, 45% of patients exhibited at least one alteration in at least one normal urothelium sample. Recurrence analysis resulted in 16 hotspots composed of either gains and copy number neutral loss of heterozygosity (CN-LOH) or deletions and CN-LOH, encompassing well-known and new BLCA cancer driver genes. Conservative assessment of LOY showed 29%, 27% and 18% of LOY-cells in tumors, blood and normal urothelium, respectively. We provide a proof of principle that our approach can characterize the earliest alterations preconditioning normal urothelium to BLCA development. Frequent LOY in blood and urothelium-derived tissues suggest its involvement in BLCA.

Background

Antimicrobial resistance (AMR) is a One Health issue and a major threat to animal and human health. Antibiotic use (ABU) drives AMR development, and several hotspots for ABU, and AMR, in livestock have been identified in Southeast Asia, including Vietnam. There are often multiple drivers of ABU at farms, and to identify all of them there is a need to look beyond farm level.

Objectives

The overall aim of this study was to identify routines and/or competencies, related to antibiotic sales, among veterinary drug shop workers that may be improved in order to decrease the medically non-rational use of antibiotics in livestock production.

Methods

A questionnaire-based survey was conducted at 50 veterinary drug shops in northern Vietnam.

Results

Results showed high education and knowledge levels. According to the respondents, antibiotic treatment advice was almost always provided to the farmers, and the recommended treatment was most commonly based on recommendations for the specific disease. However, farmers had almost never had their animals properly diagnosed. Antibiotics were the most sold drug category, penicillins being the most common. Several broad-spectrum antibiotics were also quite frequently sold. Further, >50% of respondents recommended antibiotics for disease prevention.

Conclusions

Even though education and knowledge levels might be high, several challenges can prevent drug shop workers from contributing to more prudent ABU at farms, for example, lack of proper diagnosis, commercial interests and individual farmer motives, often in combination with poor compliance to regulations.