Lactate metabolism promotes in vivo fitness during Acinetobacter baumannii infection.

Acinetobacter baumannii is one of the most prevalent causes of nosocomial infections worldwide. However, a paucity of information exists regarding the connection between metabolic capacity and in vivo bacterial fitness. Elevated lactate is a key marker of severe sepsis. We have previously shown that the putative A. baumannii lactate permease gene, lldP, is upregulated during in vivo infection. Here, we confirm that lldP expression is upregulated in three A. baumannii strains during a mammalian systemic infection. Utilising a transposon mutant disrupted for lldP in the contemporary clinical strain AB5075-UW, and a complemented strain, we confirmed its role in the in vitro utilisation of l-(+)-lactate. Furthermore, disruption of the lactate metabolism pathway resulted in reduced bacterial fitness during an in vivo systemic murine competition assay. The disruption of lldP had no impact on the susceptibility of this strain to complement mediated killing by healthy human serum. However, growth in biologically relevant concentrations of lactate observed during severe sepsis, led to bacterial tolerance to killing by healthy human blood, a phenotype that was abolished in the lldP mutant. This study highlights the importance of the lactate metabolism pathway for survival and growth of A. baumannii during infection.

Multi-Locus Microsatellite Typing of Colonising and Invasive Aspergillus fumigatus Isolates from Patients Post Lung Transplantation and with Chronic Lung Disease.

Aspergillus fumigatus can cause different clinical manifestations/phenotypes in lung transplant (LTx) recipients and patients with chronic respiratory diseases. It can also precipitate chronic lung allograft dysfunction (CLAD) in LTx recipients. Many host factors have been linked with the severity of A. fumigatus infection, but little is known about the contribution of different A. fumigatus strains to the development of different phenotypes and CLAD. We used multi-locus microsatellite typing (MLMT) to determine if there is a relationship between strain (i.e., genotype) and phenotype in 60 patients post LTx or with chronic respiratory disease across two time periods (1 November 2006-31 March 2009 and 1 November 2015-30 June 2017). The MLMT (STRAf) assay was highly discriminatory (Simpson's diversity index of 0.9819-0.9942) with no dominant strain detected. No specific genotype-phenotype link was detected, but several clusters and related strains were associated with invasive aspergillosis (IA) and colonisation in the absence of CLAD. Host factors were linked to clinical phenotypes, with prior lymphopenia significantly more common in IA cases as compared with A. fumigatus-colonised patients (12/16 [75%] vs. 13/36 [36.1%]; p = 0.01), and prior Staphylococcus aureus infection was a significant risk factor for the development of IA (odds ratio 13.8; 95% confidence interval [2.01-279.23]). A trend toward a greater incidence of CMV reactivation post-A. fumigatus isolation was observed (0 vs. 5; p = 0.06) in LTx recipients. Further research is required to determine the pathogenicity and immunogenicity of specific A. fumigatus strains.

Genomic investigation of multispecies and multivariant blaNDM outbreak reveals key role of horizontal plasmid transmission.

OBJECTIVES: New Delhi metallo-ß-lactamases (NDMs) are major contributors to the spread of carbapenem resistance globally. In Australia, NDMs were previously associated with international travel, but from 2019 we noted increasing incidence of NDM-positive clinical isolates. We investigated the clinical and genomic epidemiology of NDM carriage at a tertiary-care Australian hospital from 2016 to 2021. METHODS: We identified 49 patients with 84 NDM-carrying isolates in an institutional database, and we collected clinical data from electronic medical record. Short- and long-read whole genome sequencing was performed on all isolates. Completed genome assemblies were used to assess the genetic setting of blaNDM genes and to compare NDM plasmids. RESULTS: Of 49 patients, 38 (78%) were identified in 2019-2021 and only 11 (29%) of 38 reported prior travel, compared with 9 (82%) of 11 in 2016-2018 (P = .037). In patients with NDM infection, the crude 7-day mortality rate was 0% and the 30-day mortality rate was 14% (2 of 14 patients). NDMs were noted in 41 bacterial strains (ie, species and sequence type combinations). Across 13 plasmid groups, 4 NDM variants were detected: blaNDM-1, blaNDM-4, blaNDM-5, and blaNDM-7. We noted a change from a diverse NDM plasmid repertoire in 2016-2018 to the emergence of conserved blaNDM-1 IncN and blaNDM-7 IncX3 epidemic plasmids, with interstrain spread in 2019-2021. These plasmids were noted in 19 (50%) of 38 patients and 35 (51%) of 68 genomes in 2019-2021. CONCLUSIONS: Increased NDM case numbers were due to local circulation of 2 epidemic plasmids with extensive interstrain transfer. Our findings underscore the challenges of outbreak detection when horizontal transmission of plasmids is the primary mode of spread.

Pilot study to evaluate the need and implementation of a multifaceted nurse-led antimicrobial stewardship intervention in residential aged care.

Objectives: To evaluate the need and feasibility of a nurse-led antimicrobial stewardship (AMS) programme in two Australian residential aged care homes (RACHs) to inform a stepped-wedged, cluster randomized controlled trial (SW-cRCT). Methods: A mixed-methods pilot study of a nurse-led AMS programme was performed in two RACHs in Victoria, Australia (July-December 2019). The AMS programme comprised education, infection assessment and management guidelines, and documentation to support appropriate antimicrobial use in urinary, lower respiratory and skin/soft tissue infections. The programme was implemented over three phases: (i) pre-implementation education and integration (1 month); (ii) implementation of the intervention (3 months); and (iii) post-intervention evaluation (1 month). Baseline RACH and resident data and weekly infection and antimicrobial usage were collected and analysed descriptively to evaluate the need for AMS strategies. Feedback on intervention resources and implementation barriers were identified from semi-structured interviews, an online staff questionnaire and researcher field notes. Results: Six key barriers to implementation of the intervention were identified and used to refine the intervention: aged care staffing and capacity; access to education; resistance to practice change; role of staff in AMS; leadership and ownership of the intervention at the RACH and organization level; and family expectations. A total of 61 antimicrobials were prescribed for 40 residents over the 3 month intervention. Overall, 48% of antibiotics did not meet minimum criteria for appropriate initiation (respiratory: 73%; urinary: 54%; skin/soft tissue: 0%). Conclusions: Several barriers and opportunities to improve implementation of AMS in RACHs were identified. Findings were used to inform a revised intervention to be evaluated in a larger SW-cRCT.

Clinical and economic impact of ventricular assist device infections: a real-world claims analysis.

BACKGROUND: VAD therapy has revolutionized the treatment of end-stage heart failure, but infections remain an important complication. The objective of this study was to characterize the clinical and economic impacts of VAD-specific infections. METHODS: A retrospective analysis of a United States claims database identified members ≥ 18 years with a claim for a VAD implant procedure, at least 6 months of pre-implant baseline data, and 12 months of follow-up between 1 June 2016 and 31 December 2019. Cumulative incidence of infection was calculated. Infection and non-infection cohorts were compared regarding mortality, healthcare utilization, and total cost. Regression models were used to identify risk factors associated with infections and mortality. RESULTS: A total of 2,259 patients with a VAD implant were included, with 369 experiencing infection (12-month cumulative incidence 16.1%). Patients with infection were 2.1 times more likely to die (p < 0.001, 95% CI [1.5-2.9]). The mean 12-month total cost per US patient was $354,339 for the non-infection cohort and $397,546 for the infection cohort, a difference of $43,207 (p < 0.0001). CONCLUSIONS: VAD infections were associated with higher mortality, more healthcare utilization, and higher total cost. Strategies to minimize VAD-specific infections could lead to improved clinical and economic outcomes.

PmxPred: A data-driven approach for the identification of active polymyxin analogues against gram-negative bacteria.

The multidrug-resistant Gram-negative bacteria has evolved into a worldwide threat to human health; over recent decades, polymyxins have re-emerged in clinical practice due to their high activity against multidrug-resistant bacteria. Nevertheless, the nephrotoxicity and neurotoxicity of polymyxins seriously hinder their practical use in the clinic. Based on the quantitative structure-activity relationship (QSAR), analogue design is an efficient strategy for discovering biologically active compounds with fewer adverse effects. To accelerate the polymyxin analogues discovery process and find the polymyxin analogues with high antimicrobial activity against Gram-negative bacteria, here we developed PmxPred, a GCN and catBoost-based machine learning framework. The RDKit descriptors were used for the molecule and residues representation, and the ensemble learning model was utilized for the antimicrobial activity prediction. This framework was trained and evaluated on multiple Gram-negative bacteria datasets, including Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and a general Gram-negative bacteria dataset achieving an AUROC of 0.857, 0.880, 0.756, 0.895 and 0.865 on the independent test, respectively. PmxPred outperformed the transfer learning method that trained on 10 million molecules. We interpreted our model well-trained model by analysing the importance of global and residue features. Overall, PmxPred provides a powerful additional tool for predicting active polymyxin analogues, and holds the potential elucidate the mechanisms underlying the antimicrobial activity of polymyxins. The source code is publicly available on GitHub (https://github.com/yanwu20/PmxPred).

Polysaccharide-Targeting Lipid Nanoparticles to Kill Gram-Negative Bacteria.

The rapid increase and spread of Gram-negative bacteria resistant to many or all existing treatments threaten a return to the preantibiotic era. The presence of bacterial polysaccharides that impede the penetration of many antimicrobials and protect them from the innate immune system contributes to resistance and pathogenicity. No currently approved antibiotics target the polysaccharide regions of microbes. Here, describe monolaurin-based niosomes, the first lipid nanoparticles that can eliminate bacterial polysaccharides from hypervirulent Klebsiella pneumoniae, are described. Their combination with polymyxin B shows no cytotoxicity in vitro and is highly effective in combating K. pneumoniae infection in vivo. Comprehensive mechanistic studies have revealed that antimicrobial activity proceeds via a multimodal mechanism. Initially, lipid nanoparticles disrupt polysaccharides, then outer and inner membranes are destabilized and destroyed by polymyxin B, resulting in synergistic cell lysis. This novel lipidic nanoparticle system shows tremendous promise as a highly effective antimicrobial treatment targeting multidrug-resistant Gram-negative pathogens.

Candida causes recurrent vulvovaginal candidiasis by forming morphologically disparate biofilms on the human vaginal epithelium.

Background: Recurrent vulvovaginal candidiasis (RVVC) is a recalcitrant medical condition that affects many women of reproductive age. The importance of biofilm formation by Candida in RVVC has been recently questioned. This study aimed to elucidate the fundamental growth modes of Candida in the vagina of patients with RVVC or sporadic vulvovaginal candidiasis (VVC) and to assess their roles in the persistence of RVVC. Methods: Vaginal tissues were sampled from twelve patients clinically and microbiologically diagnosed as RVVC or VVC at a post-antifungal-treatment and asymptomatic period. High-resolution scanning electron microscopy, fluorescence in situ hybridization in combination with Candida-specific 18S rRNA probes and viable fungal burden were used to qualitatively and quantitatively evaluate Candida growth in the human vagina. The presence of Candida biofilm extracellular polymeric substances was examined using confocal laser scanning microscopy and biopsy sections pre-stained with Concanavalin A. Histopathological analysis was carried out on infected vaginal tissues stained with hematoxylin and eosin. Lastly, the susceptibility of epithelium-associated Candida biofilms to fluconazole at the peak serum concentration was evaluated. Results: Candida species grew on the vaginal epithelium of RVVC patients as morphologically disparate biofilms including monolayers, microcolonies, and macro-colonies, in addition to sporadic adherent cells. Candida biofilm growth on the vaginal epithelium was associated with mild lymphocytic infiltration of the vaginal mucosa. These epithelium-based Candida biofilms presented an important characteristic contributing to the persistence of RVVC that is the high tolerance to fluconazole. Conclusions: In summary, our study provides direct evidence to support the presence of Candida biofilms in RVVC and an important role of biofilm formation in disease persistence.

Virulence attributes of successful methicillin-resistant Staphylococcus aureus lineages.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of severe and often fatal infections. MRSA epidemics have occurred in waves, whereby a previously successful lineage has been replaced by a more fit and better adapted lineage. Selection pressures in both hospital and community settings are not uniform across the globe, which has resulted in geographically distinct epidemiology. This review focuses on the mechanisms that trigger the establishment and maintenance of current, dominant MRSA lineages across the globe. While the important role of antibiotic resistance will be mentioned throughout, factors which influence the capacity of S. aureus to colonize and cause disease within a host will be the primary focus of this review. We show that while MRSA possesses a diverse arsenal of toxins including alpha-toxin, the success of a lineage involves more than just producing toxins that damage the host. Success is often attributed to the acquisition or loss of genetic elements involved in colonization and niche adaptation such as the arginine catabolic mobile element, as well as the activity of regulatory systems, and shift metabolism accordingly (e.g., the accessory genome regulator, agr). Understanding exactly how specific MRSA clones cause prolonged epidemics may reveal targets for therapies, whereby both core (e.g., the alpha toxin) and acquired virulence factors (e.g., the Panton-Valentine leukocidin) may be nullified using anti-virulence strategies.

A genomic survey of Clostridioides difficile isolates from hospitalized patients in Melbourne, Australia.

IMPORTANCE: There has been a decrease in healthcare-associated Clostridioides difficile infection in Australia, but an increase in the genetic diversity of infecting strains, and an increase in community-associated cases. Here, we studied the genetic relatedness of C. difficile isolated from patients at a major hospital in Melbourne, Australia. Diverse ribotypes were detected, including those associated with community and environmental sources. Some types of isolates were more likely to carry antimicrobial resistance determinants, and many of these were associated with mobile genetic elements. These results correlate with those of other recent investigations, supporting the observed increase in genetic diversity and prevalence of community-associated C. difficile, and consequently the importance of sources of transmission other than symptomatic patients. Thus, they reinforce the importance of surveillance for in both hospital and community settings, including asymptomatic carriage, food, animals, and other environmental sources to identify and circumvent important sources of C. difficile transmission.

Clinical Staphylococcus aureus inhibits human T-cell activity through interaction with the PD-1 receptor.

IMPORTANCE: Therapies that target and aid the host immune defense to repel cancer cells or invading pathogens are rapidly emerging. Antibiotic resistance is among the largest threats to human health globally. Staphylococcus aureus (S. aureus) is the most common bacterial infection, and it poses a challenge to the healthcare system due to its significant ability to develop resistance toward current available therapies. In long-term infections, S. aureus further adapt to avoid clearance by the host immune defense. In this study, we discover a new interaction that allows S. aureus to avoid elimination by the immune system, which likely supports its persistence in the host. Moreover, we find that blocking the specific receptor (PD-1) using antibodies significantly relieves the S. aureus-imposed inhibition. Our findings suggest that therapeutically targeting PD-1 is a possible future strategy for treating certain antibiotic-resistant staphylococcal infections.

EHR-QC: A streamlined pipeline for automated electronic health records standardisation and preprocessing to predict clinical outcomes.

The adoption of electronic health records (EHRs) has created opportunities to analyse historical data for predicting clinical outcomes and improving patient care. However, non-standardised data representations and anomalies pose major challenges to the use of EHRs in digital health research. To address these challenges, we have developed EHR-QC, a tool comprising two modules: the data standardisation module and the preprocessing module. The data standardisation module migrates source EHR data to a standard format using advanced concept mapping techniques, surpassing expert curation in benchmarking analysis. The preprocessing module includes several functions designed specifically to handle healthcare data subtleties. We provide automated detection of data anomalies and solutions to handle those anomalies. We believe that the development and adoption of tools like EHR-QC is critical for advancing digital health. Our ultimate goal is to accelerate clinical research by enabling rapid experimentation with data-driven observational research to generate robust, generalisable biomedical knowledge.

Genomic dissection of endemic carbapenem resistance reveals metallo-beta-lactamase dissemination through clonal, plasmid and integron transfer.

Infections caused by metallo-beta-lactamase-producing organisms (MBLs) are a global health threat. Our understanding of transmission dynamics and how MBLs establish endemicity remains limited. We analysed two decades of blaIMP-4 evolution in a hospital using sequence data from 270 clinical and environmental isolates (including 169 completed genomes) and identified the blaIMP-4 gene across 7 Gram-negative genera, 68 bacterial strains and 7 distinct plasmid types. We showed how an initial multi-species outbreak of conserved IncC plasmids (95 genomes across 37 strains) allowed endemicity to be established through the ability of blaIMP-4 to disseminate in successful strain-genetic setting pairs we termed propagators, in particular Serratia marcescens and Enterobacter hormaechei. From this reservoir, blaIMP-4 persisted through diversification of genetic settings that resulted from transfer of blaIMP-4 plasmids between bacterial hosts and of the integron carrying blaIMP-4 between plasmids. Our findings provide a framework for understanding endemicity and spread of MBLs and may have broader applicability to other carbapenemase-producing organisms.

The clinical application of beta-lactam antibiotic therapeutic drug monitoring in the critical care setting.

Critically ill patients have increased variability in beta-lactam antibiotic (beta-lactam) exposure due to alterations in their volume of distribution and elimination. Therapeutic drug monitoring (TDM) of beta-lactams, as a dose optimization and individualization tool, has been recommended to overcome this variability in exposure. Despite its potential benefit, only a few centres worldwide perform beta-lactam TDM. An important reason for the low uptake is that the evidence for clinical benefits of beta-lactam TDM is not well established. TDM also requires the availability of specific infrastructure, knowledge and expertise. Observational studies and systematic reviews have demonstrated that TDM leads to an improvement in achieving target concentrations, a reduction in potentially toxic concentrations and improvement of clinical and microbiological outcomes. However, a small number of randomized controlled trials have not shown a mortality benefit. Opportunities for improved study design are apparent, as existing studies are limited by their inclusion of heterogeneous patient populations, including patients that may not even have infection, small sample size, variability in the types of beta-lactams included, infections caused by highly susceptible bacteria, and varied sampling, analytical and dosing algorithm methods. Here we review the fundamentals of beta-lactam TDM in critically ill patients, the existing clinical evidence and the practical aspects involved in beta-lactam TDM implementation.

Characterisation of key genotypic and phenotypic traits of clinical cystic fibrosis Staphylococcus aureus isolates.

Introduction. One third of people with CF in the UK are co-infected by both Staphylococcus aureus and Pseudomonas aeruginosa. Chronic bacterial infection in CF contributes to the gradual destruction of lung tissue, and eventually respiratory failure in this group.Gap Statement. The contribution of S. aureus to cystic fibrosis (CF) lung decline in the presence or absence of P. aeruginosa is unclear. Defining the molecular and phenotypic characteristics of a range of S. aureus clinical isolates will help further understand its pathogenic capabilities.Aim. Our objective was to use molecular and phenotypic tools to characterise twenty-five clinical S. aureus isolates collected from mono- and coinfection with P. aeruginosa from people with CF at the Royal Victoria Infirmary, Newcastle upon Tyne.Methodology. Genomic DNA was extracted and sequenced. Multilocus sequence typing was used to construct phylogeny from the seven housekeeping genes. A pangenome was calculated using Roary, and cluster of Orthologous groups were assigned using eggNOG-mapper which were used to determine differences within core, accessory, and unique genomes. Characterisation of sequence type, clonal complex, agr and spa types was carried out using PubMLST, eBURST, AgrVATE and spaTyper, respectively. Antibiotic resistance was determined using Kirby-Bauer disc diffusion tests. Phenotypic testing of haemolysis was carried out using ovine red blood cell agar plates and mucoid phenotypes visualised using Congo red agar.Results. Clinical strains clustered closely based on agr type, sequence type and clonal complex. COG analysis revealed statistically significant enrichment of COG families between core, accessory and unique pangenome groups. The unique genome was significantly enriched for replication, recombination and repair, and defence mechanisms. The presence of known virulence genes and toxins were high within this group, and unique genes were identified in 11 strains. Strains which were isolated from the same patient all surpassed average nucleotide identity thresholds, however, differed in phenotypic traits. Antimicrobial resistance to macrolides was significantly higher in the coinfection group.Conclusion. There is huge variation in genetic and phenotypic capabilities of S. aureus strains. Further studies on how these may differ in relation to other species in the CF lung may give insight into inter-species interactions.

Antimicrobial susceptibility of ceftolozane-tazobactam against multidrug-resistant Pseudomonas aeruginosa isolates from Melbourne, Australia.

We collected 163 clinical Pseudomonas aeruginosa isolates at a tertiary hospital specialising in adult cystic fibrosis (CF) and lung transplantation (LTx) in Melbourne, Australia, to explore the activity of ceftolozane-tazobactam (C/T) in populations at high-risk for antimicrobial resistance. Of these, 144 (88.3%) were collected from sputum, and 19 (11.7%) from bronchoalveolar lavage. Most (85.3%) were derived from patients with cystic fibrosis and included a subset of patients that had undergone LTx. These isolates were tested against 11 antibiotics, including C/T, using Sensititre plates for broth microdilution (BMD) testing. Sixty (36.8%) isolates were classified as multidrug resistant (MDR) and 32 (19.6%) were extensively drug resistant (XDR). Overall, 133/163 (81.6%) isolates were susceptible to C/T. For MDR and XDR isolates, 88.3% and 28.1% were C/T susceptible, respectively. Among the non-MDR/XDR isolates, 100% remained susceptible to C/T. Comparisons of C/T susceptibility were made using BioMérieux Etests and Liofilchem MIC test strips (MTS). Categorical agreement to BMD was >93% for both test strips, but essential agreement to BMD was slightly higher with Etest (89.0%) compared to Liofilchem (74.8%). In conclusion, C/T retained activity against most MDR and over a quarter of XDR P. aeruginosa isolates from complex patients with CF and post-LTx.

Understanding patient and healthcare worker experiences and perspectives of multidrug-resistant organisms.

Objectives: Transmission of MDR organisms (MROs) such as carbapenemase-producing Enterobacteriaceae (CPE) and VRE in healthcare facilities is a major issue globally. Knowledge gaps exist, including the impact of these microorganisms on patients, and healthcare worker understanding of infection control approaches for MROs. This study aimed to explore patient and healthcare worker experiences and perspectives of MROs. Methods: A sequential exploratory mixed-methods study was performed at a large metropolitan acute and subacute hospital. This involved semi-structured face-to-face interviews with patients with confirmed MROs to explore their understanding of these microorganisms and perceptions of their time in hospital. Healthcare workers participated in an online survey about their understanding of MROs and the care of patients with these microorganisms. Qualitative data were analysed using the COM-B framework, and were triangulated with the descriptive quantitative analysis. Results: The overarching theme from the triangulated data was uncertainty amongst both patients and staff about MROs. Insufficient explanations from staff left patients lacking a proper understanding of their diagnosis, and patients felt that staff did not always follow isolation protocols. Staff felt they did not receive enough education on MROs. However, patients felt that the overall care they received was very good, and most valued the privacy gained from being in isolation. Conclusions: This study demonstrates that there is a need to focus on new strategies of communication with patients and staff education to improve understanding of MROs and increase adherence to protocols.

iAMPCN: a deep-learning approach for identifying antimicrobial peptides and their functional activities.

Antimicrobial peptides (AMPs) are short peptides that play crucial roles in diverse biological processes and have various functional activities against target organisms. Due to the abuse of chemical antibiotics and microbial pathogens' increasing resistance to antibiotics, AMPs have the potential to be alternatives to antibiotics. As such, the identification of AMPs has become a widely discussed topic. A variety of computational approaches have been developed to identify AMPs based on machine learning algorithms. However, most of them are not capable of predicting the functional activities of AMPs, and those predictors that can specify activities only focus on a few of them. In this study, we first surveyed 10 predictors that can identify AMPs and their functional activities in terms of the features they employed and the algorithms they utilized. Then, we constructed comprehensive AMP datasets and proposed a new deep learning-based framework, iAMPCN (identification of AMPs based on CNNs), to identify AMPs and their related 22 functional activities. Our experiments demonstrate that iAMPCN significantly improved the prediction performance of AMPs and their corresponding functional activities based on four types of sequence features. Benchmarking experiments on the independent test datasets showed that iAMPCN outperformed a number of state-of-the-art approaches for predicting AMPs and their functional activities. Furthermore, we analyzed the amino acid preferences of different AMP activities and evaluated the model on datasets of varying sequence redundancy thresholds. To facilitate the community-wide identification of AMPs and their corresponding functional types, we have made the source codes of iAMPCN publicly available at https://github.com/joy50706/iAMPCN/tree/master. We anticipate that iAMPCN can be explored as a valuable tool for identifying potential AMPs with specific functional activities for further experimental validation.

Anti-infective characteristics of a new Carbothane ventricular assist device driveline.

Objectives: Driveline infections are a major complication of ventricular assist device (VAD) therapy. A newly introduced Carbothane driveline has preliminarily demonstrated anti-infective potential against driveline infections. This study aimed to comprehensively assess the anti-biofilm capability of the Carbothane driveline and explore its physicochemical characteristics. Methods: We assessed the Carbothane driveline against biofilm formation of leading microorganisms causing VAD driveline infections, including Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa and Candida albicans, using novel in vitro biofilm assays mimicking different infection micro-environments. The importance of physicochemical properties of the Carbothane driveline in microorganism-device interactions were analyzed, particularly focusing on the surface chemistry. The role of micro-gaps in driveline tunnels on biofilm migration was also examined. Results: All organisms were able to attach to the smooth and velour sections of the Carbothane driveline. Early microbial adherence, at least for S. aureus and S. epidermidis, did not proceed to the formation of mature biofilms in a drip-flow biofilm reactor mimicking the driveline exit site environment. The presence of a driveline tunnel however, promoted staphylococcal biofilm formation on the Carbothane driveline. Physicochemical analysis of the Carbothane driveline revealed surface characteristics that may have contributed to its anti-biofilm activity, such as the aliphatic nature of its surface. The presence of micro-gaps in the tunnel facilitated biofilm migration of the studied bacterial species. Conclusion: This study provides experimental evidence to support the anti-biofilm activity of the Carbothane driveline and uncovered specific physicochemical features that may explain its ability to inhibit biofilm formation.