Biology subjects

Bandara, A.

Publications and source records attributed to Bandara, A..

5 recordsLinked to original sources

bioRxiv2026

Impairment of bacteriophage activity in blood: a case study revealing constraints in phage isolation and translation

BackgroundPhage therapy is increasingly considered a promising alternative for treating multidrug-resistant (MDR) infections. However, its clinical application remains limited by challenges in isolating effective phages against resistant clinical strains and by the limited ability of in vitro assays to predict performance in real biological environments. While biological matrices are known to influence phage activity, these effects are not well characterised. MethodsA phage-resistant Pseudomonas aeruginosa isolate from a patient with recurrent MDR urinary tract infection was used as the model organism. Conventional isolation methods failed to recover effective phages, leading to the development of TEASER-i (Transient EDTA- and Ion-Assisted Sequential Enrichment & Recovery). Recovered phages were characterised using adsorption assays, one-step growth kinetics, and time-kill experiments. Their antibacterial activity was evaluated both in vitro and in ex vivo human matrices (whole blood, serum, plasma, and urine). Phage efficacy was quantified using maximum log reduction (Emax), area under the curve (AUC), and phage-to-bacteria ratio (PBR). ResultsA novel TEASER-i method optimised for difficult-to-treat Gram-negative infections, enabled recovery of a functionally effective Osewage-derived P. aeruginosa phage, which outperformed a Ourine-derived P. aeruginosa phage that showed slower replication and lower burst size. Phage activity varied significantly in blood, serum, and plasma. Urine supported the most sustained antibacterial effect. In many cases, early bacterial reduction was followed by regrowth. Sustained activity was associated with maintenance of favourable PBR values, while negative PBR corresponded to treatment failure. At 96 h, only two conditions maintained favourable phage load (log 10 PBR > 0): the S. aureus phage in urine (+1.66) and the sewage-derived P. aeruginosa phage in serum (+1.32). ConclusionsPhage efficacy depends not only on intrinsic lytic capacity but also on the ability to persist and amplify within specific biological environments. Conventional isolation and in vitro screening may therefore overestimate therapeutic potential. Combining optimised isolation strategies with ex vivo evaluation provides a more realistic framework for phage selection and clinical translation.

microbiology↗

bioRxiv2025

Natural and patient-derived mutations in BK polyomavirus VP1 reveal structural determinants of BC-loop dependent antibody escape

BK polyomavirus (BKPyV) reactivation poses a serious threat to both renal and hematopoietic transplant outcomes, with no approved antiviral therapies. Neutralizing monoclonal antibodies targeting the viral capsid protein VP1 have recently advanced into clinical development, driven by promising in vitro potency. Since naturally elicited neutralizing antibodies arise following infection, we examined whether host immune pressure may have shaped the VP1 surface, with implications for therapeutic application. To probe the impact of VP1 coevolution on therapeutic engagement, we evaluated two neutralizing antibody formats: the clinical-stage monoclonal antibody 319C07 and nanobodies VHH16 and VHH17. Our X-ray crystallographic analysis of the VP1 pentameric core in complex with antibody fragments showed a convergent receptor- mimetic engagement of the sialic acid-binding cleft, mapping to a shared epitope centered around the VP1 BC-loop. Analysis of over 900 BKPyV VP1 sequences from public databases, combined with longitudinal VP1 sequence data from transplant recipients, highlighted widespread pre-existing diversity at the BC-loop contact sites, underscoring the evolutionary adaptability of BKPyV at this antigenic surface that may compromise therapeutic recognition. Through integrated mutagenesis and binding analyses, we show that circulating BC-loop mutations, including single amino acid substitutions, are sufficient to abrogate 319C07 binding and neutralization. Nanobodies VHH16 and VHH17, selected for their compact size and potential to access intrarenal sites of BKPyV replication, also exhibited complete loss of binding across multiple clinically observed VP1 variants. Our findings may offer a mechanistic framework to interpret the translational gap between in vitro neutralization and clinical efficacy of these biologics. As part of ongoing efforts to target VP1, we demonstrate, for the first time, small molecules with nanomolar to sub-nanomolar affinities against BKPyV VP1 variants including potent binding to JCPyV VP1. Our work paves a path towards a new class of antiviral strategies that could block replication at intracellular replication sites, provide a higher genetic barrier to resistance and the potential to address both viral reactivation and persistent, high- burden viremia in transplant settings.

microbiology↗

bioRxiv2025

Exploring the kinetics and mechanism of phase separation in ternary lipid mixtures containing APP C99 using atomistic vs coarse-grained MD simulations

The phase separation of lipid bilayers, composed of mixtures of saturated and unsaturated lipids and cholesterol, is a topic of fundamental importance in membrane biophysics and cell biology. The formation of lipid domains, including liquid-disordered domains enriched in unsaturated lipids and liquid-ordered domains enriched in saturated lipids and cholesterol is believed to be essential to the function of many membrane proteins. Experiment, theory, and simulation have been used to develop a general understanding of the thermodynamic driving forces underlying phase separation in ternary and quaternary lipid mixtures. However, the kinetics of early events in lipid phase separation in the presence of transmembrane proteins remain relatively understudied. Using large-scale all-atom and coarse-grained simulations, we explore the kinetics and phase separation of ternary lipid mixtures of saturated lipid, unsaturated lipid, and cholesterol. Order parameters employed in the Cahn-Hilliard theory provide insight into the kinetics and mechanism of lipid phase separation. We observe three distinct time regimes in the phase separation process: a shorter time exponential phase followed by a power law phase followed by a longer time plateau phase. Comparison of lipid, protein and lipid-protein dynamics between all-atom and coarse-grained models identifies both quantitative and qualitative differences and similarities in the phase separation kinetics. Moreover, timescaling of dynamics of AA and CG simulation yields a similar kinetic mechanism of phase separation. The findings of this study elucidate fundamental aspects of membrane biophysics and the ongoing efforts to define the role of lipid rafts in the structure and function of cellular membrane.

biophysics↗

bioRxiv2024

A Novel Magnetic Field Device: Effects of Magnetic Fields on Planktonic Yeasts and Fungal Mats

Microorganisms evolved within the geomagnetic field and can be affected by magnetic field exposure. However, the mechanisms underlying many magnetic phenomena in microbes remain to be elucidated. We develop a 3D-printed magnetic field exposure device to perform experiments on microbes. This device is designed in AutoCAD, modeled in COMSOL, and validated using a Gaussmeter. Using the magnetic field exposure device, we perform static magnetic field experiments on different strains of the budding yeast Saccharomyces cerevisiae. We find that static magnetic field exposure slows the spatially-structured expansion of yeast mats that expands in two dimensions, but not yeast mats that expand in three dimensions, across the surface of semi-solid media. We also find that magnetic fields do not affect the growth of yeast cells in well-mixed liquid media. This study provides a novel device for magnetic field exposure experiments on microorganisms and advances our understanding of the effects of magnetic fields on fungi. Why it mattersMicroorganisms have evolved to function, survive, and reproduce in Earths magnetic field. However, the mechanisms underlying magnetic phenomena in microorganisms are unknown. This is especially true for fungi, which are important microorganisms for microbiological research, industrial application, and infectious disease. To elucidate mechanisms driving magnetic phenomena, we need devices to perform controlled experiments in a variety of conditions. We develop a 3D-printed magnetic field exposure device using computer-aided design, physics modeling software, and a magnetometer. Using this novel magnetic field device, we discover that magnetic fields can slow the growth of yeast on agar plates, but that magnetic fields do not affect the growth of yeast in liquid media.

biophysics↗

bioRxiv2020

The Effect Of Famotidine On SARS-CoV-2 Proteases And Virus Replication

The lack of coronavirus-specific antiviral drugs has instigated multiple drug repurposing studies to redirect previously approved medicines for the treatment of SARS-CoV-2, the coronavirus behind the ongoing COVID-19 pandemic. A recent, large-scale, retrospective clinical study showed that famotidine, when administered at a high dose to hospitalized COVID-19 patients, reduced the rates of intubation and mortality. A separate, patient-reported study associated famotidine use with improvements in mild to moderate symptoms such as cough and shortness of breath. While a prospective, multi-center clinical study is ongoing, two parallel in silico studies have proposed one of the two SARS-CoV-2 proteases, 3CLpro or PLpro, as potential molecular targets of famotidine activity; however, this remains to be experimentally validated. In this report, we systematically analyzed the effect of famotidine on viral proteases and virus replication. Leveraging a series of biophysical and enzymatic assays, we show that famotidine neither binds with nor inhibits the functions of 3CLpro and PLpro. Similarly, no direct antiviral activity of famotidine was observed at concentrations of up to 200 M, when tested against SARS-CoV-2 in two different cell lines, including a human cell line originating from lungs, a primary target of COVID-19. These results rule out famotidine as a direct-acting inhibitor of SARS-CoV-2 replication and warrant further investigation of its molecular mechanism of action in the context of COVID-19.

pharmacology and toxicology↗