Biology subjects

Fanning, S.

Publications and source records attributed to Fanning, S..

7 recordsLinked to original sources

bioRxiv2026

Glyphosate, a herbicide, and fosfomycin, an antibiotic in clinical use- evidence of common selectable genotypes

The emergence of antimicrobial resistance (AMR) is increasingly linked to metabolic adaptation, yet the evolutionary routes underlying cross-resistance between structurally related compounds remain poorly understood. Here, whole genome sequencing (WGS) was used to analyse Klebsiella pneumoniae mutants evolved under sub-lethal glyphosate (GLP) or fosfomycin (FOS) exposure to determine how these stresses shape resistance and physiology. Sub-lethal GLP exposure increased FOS resistance, demonstrating cross-resistance between the two phosphonates. FOS-evolved mutants achieved high-level resistance through the accumulation of multiple mutations affecting the antibiotic target MurA, transport systems, and global metabolic regulation, producing a layered FOS resistance phenotype. In contrast, GLP-evolved mutants acquired similar functional classes of mutations but exhibited lower baseline FOS resistance, suggesting trade-offs between resistance and metabolic fitness. Further, analysis of FOS-evolved and GLP-evolved mutants across known bacterial GLP resistance mechanisms demonstrated a strong overlap. Comparative genomic analysis revealed a small, recurrent set of genes under selection in both evolutionary trajectories, with identical genomic loci repeatedly targeted, consistent with convergent evolution. Many of these changes were linked to central metabolism, redox balance, and cell surface regulation. For some isolates, a hypermutator phenotype was necessary to offset the potentially lethal effects of primary-target mutations through compensatory genomic adaptation. In conclusion, GLP and FOS select for shared adaptive networks that couple metabolic rewiring with AMR, revealing cross-resistance as an emergent property of global physiological reprogramming and providing mechanistic insight into ecological models of co-selection in environmental systems. Importance statementGlyphosate is a herbicide in current use worldwide. Its impact on the susceptibility of bacteria to antibiotics, remains to be described. This manuscript details a critical phenotypic and genomic analysis of shared resistance mechanisms between glyphosate and the antibiotic fosfomycin. Using Klebsiella pneumoniae, a zoonotic pathogen, this manuscript demonstrates that evolutionary adaptation to either compound results in a substantial overlap in gene mutations. Crucially, this manuscript shows that exposure to sub-lethal concentrations of glyphosate can increase resistance to fosfomycin. These findings reveal a link between agricultural chemical use and the emergence of cross-resistance to fosfomycin. By highlighting how environmental factors drive antimicrobial resistance, this study underscores an urgent need for revision of food safety and regulatory frameworks to protect One Health. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=185 SRC="FIGDIR/small/725383v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@d44420org.highwire.dtl.DTLVardef@1c4a5d1org.highwire.dtl.DTLVardef@1624052org.highwire.dtl.DTLVardef@16d08d4_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2025

Rational Design and Optimization of a Potent IDO1 Proteolysis Targeting Chimera (PROTAC)

Indoleamine 2,3-dioxygenase 1 (IDO1) is a potently immunosuppressive protein that inhibits antitumor immunity through both tryptophan metabolism and non-enzymatic functions. Pharmacological therapies targeting IDO1 enzyme activity have generally failed to improve the overall survival of patients with cancer. Developing new therapeutic agents that are capable of neutralizing both enzyme-and non-enzyme-derived immunosuppressive IDO1 effects is therefore of high interest. We previously described the development of a novel Proteolysis Targeting Chimera (PROTAC), NU223612, that degrades IDO1 in cultured human glioblastoma (GBM) cells, as well as in well-established brain tumors, in vivo. In this study, we rationally optimized the composition, rigidity, and linker orientation of the PROTAC structure to create NU227326, which degrades IDO1 with a DC50 of 5 nM in human GBM cells. Mechanistic studies showed that IDO1 degradation occurred through the ubiquitin-proteasome system and was sustained for at least 2 days, supporting NU227326 as a highly potent IDO1 PROTAC suitable for further studies in GBM and other human cancers.

biochemistry↗

bioRxiv2024

The Role of Alpha Synuclein in Synucleinopathy: Impact on Lipid Regulation at Mitochondria ER Membranes

The protein alpha-synuclein (Syn) plays a critical role in the pathogenesis of synucleinopathy, which includes Parkinsons disease and multiple system atrophy, and mounting evidence suggests that lipid dyshomeostasis is a critical phenotype in these neurodegenerative conditions. Previously, we identified that Syn localizes to mitochondria-associated endoplasmic reticulum membranes (MAMs), temporary functional domains containing proteins that regulate lipid metabolism, including the de novo synthesis of phosphatidylserine. In the present study, we have analyzed the lipid composition of postmortem human samples, focusing on the substantia nigra pars compacta of Parkinsons disease and controls, as well as three less affected brain regions of Parkinsons donors. To further assess synucleinopathy-related lipidome alterations, similar analyses were performed on the striatum of multiple system atrophy cases. Our data show region-and disease-specific changes in the levels of lipid species. Specifically, our data revealed alterations in the levels of specific phosphatidylserine species in brain areas most affected in Parkinsons disease. Some of these alterations, albeit to a lesser degree, are also observed multiples system atrophy. Using induced pluripotent stem cell-derived neurons, we show that Syn contributes to regulating phosphatidylserine metabolism at MAM domains, and that Syn dosage parallels the perturbation in phosphatidylserine levels. Our results support the notion that Syn pathophysiology is linked to the dysregulation of lipid homeostasis, which may contribute to the vulnerability of specific brain regions in synucleinopathy. These findings have significant therapeutic implications. Significance StatementSynucleinopathy is a complex group of neurodegenerative disorders whose causes and underlying mechanisms remain unknown. In this work, we examined synucleinopathy postmortem brain samples and patient-derived neuron models and identified the functional impairment of the mitochondrial-associated endoplasmic reticulum membrane (MAM) domain, which facilitates lipid regulation. The protein alpha-synuclein is associated with synucleinopathy and increasing levels result in the mislocalization of this protein and the disruption of MAM domains, which, in turn, results in lipid and membrane composition alterations. Specifically, we report that increased alpha-synuclein expression impairs the regulation of phosphatidylserine synthase 2 and the levels of phosphatidylserine in cellular membranes from affected cells. Our study offers mechanistic insight tying alpha-synuclein pathology and lipid dysregulation as seminal factors in synucleinopathy, which may have pathogenic and therapeutic implications.

neuroscience↗

bioRxiv2021

A study of the correlation between phenotypic antimicrobial susceptibility testing methods and the associated genotypes determined by whole genome sequencing for a collection of Escherichia coli of bovine origin

Antimicrobial resistance (AMR) has increased at an alarming pace in the recent years. Molecular-based methods such as whole genome sequencing (WGS) offer a potential alternative to the conventional labour-intensive methods traditionally used to characterise AMR phenotypes. The aim of this study was to investigate whether WGS could be used as a predictor of AMR in Escherichia coli isolates of bovine origin. Genomes of 143 E. coli cultured from cattle presenting with diarrhoea or mastitis were sequenced on an Illumina MiSeq platform. AMR genes were identified using the ResFinder and AMRFinder databases. Antimicrobial susceptibility testing by disk diffusion was performed on a panel of 10 antibiotics, covering 7 antimicrobial classes. Minimum inhibitory concentration (MIC) measurements were made using the Sensititre plate with 6 antibiotics, covering 5 antimicrobial classes. Correlation between genotype and phenotype was assessed statistically by means of a two-by-two table analysis and Cohens kappa ({kappa}) test. The overall {kappa} correlation between WGS and disk diffusion was 0.81, indicating a near perfect agreement, and the average positive predicted value was 77.4 %. Correlation for individual antimicrobial compounds varied, with five yielding near perfect agreement ({kappa} = 0.81-1.00; amoxicillin, florfenicol, gentamicin, tetracycline and trimethoprim-sulfamethoxazole), one showing substantial agreement ({kappa} = 0.65; nalidixic acid), and four showing moderate agreement ({kappa} = 0.41- 0.60). The overall {kappa} correlation between WGS and MIC was 0.55 indicating moderate agreement, and the average positive predicted value was 68.6 %. Three antibiotics yielded near perfect agreement (gentamicin, tetracycline and trimethoprim-sulfamethoxazole) and a further three showed fair agreement ({kappa} = 0.21-0.40). WGS is a useful tool that can be used for the prediction of AMR phenotypes, and correlates well with disk diffusion results. MIC measurements may be necessary for antimicrobial compounds with a high proportion of intermediately resistant isolates recorded, such as cephalothin. HighlightsO_LICulture based antimicrobial susceptibility testing is used to identify therapeutics in the treatment of clinical veterinary isolates C_LIO_LIWhole genome sequencing is increasingly adopted for surveillance, epidemiological traceback investigations, and detection of antimicrobial resistance genes C_LIO_LILittle is known in correlations between antimicrobial resistance genotypes and disk diffusion antimicrobial susceptibility testing C_LIO_LIThis study finds that whole genome sequencing is a useful predictor for antimicrobial susceptibility however, minimum inhibitory concentration measurements may still be needed for intermediately resistant isolates C_LI

microbiology↗

bioRxiv2020

Silver nanoparticles induce a triclosan like antibacterial action mechanism in multi-drug resistant Klebsiella pneumoniae

Infections associated with antimicrobial-resistant bacteria now represent a significant threat to human health using conventional therapy, necessitating the development of alternate and more effective antibacterial compounds. Silver nanoparticles (Ag NPs) have been proposed as potential antimicrobial agents to combat infections. A complete understanding of their antimicrobial activity is required before these molecules can be used in therapy. Lysozyme coated Ag NPs were synthesized and characterized by TEM-EDS, XRD, UV-vis, FTIR spectroscopy, zeta potential and oxidative potential assay. Biochemical assays and deep level transcriptional analysis using RNA sequencing were used to decipher how Ag NPs exert their antibacterial action against multi-drug resistant Klebsiella pneumoniae MGH78578. RNAseq data revealed that Ag NPs induced a triclosan-like bactericidal mechanism responsible for the inhibition of the type II fatty acid biosynthesis. Additionally, released Ag+ generated oxidative stress both extra- and intracellularly in K. pneumoniae. The data showed that triclosan-like activity and oxidative stress cumulatively underpinned the antibacterial activity of Ag NPs. This result was confirmed by the analysis of the bactericidal effect of Ag NPs against the isogenic K. pneumoniae MGH78578{Delta} soxS mutant, which exhibits a compromised oxidative stress response compared to the wild type. Silver nanoparticles induce a triclosan-like antibacterial action mechanism in multi-drug resistant K. pneumoniae. This study extends our understanding of anti-Klebsiella mechanisms associated with exposure to Ag NPs. This allowed us to model how bacteria might develop resistance against silver nanoparticles, should the latter be used in therapy.

microbiology↗

bioRxiv2020

Analysis of the oxidative stress regulon identifies soxS as a genetic target for resistance reversal in multi-drug resistant Klebsiella pneumoniae

In bacteria, the defense system deployd to counter oxidative stress is orchestrated by three transcriptional factors - SoxS, SoxR, and OxyR. Although the regulon that these factors control is known in many bacteria, similar data is not available for Klebsiella pneumoniae. To address this data gap, oxidative stress was artificially induced in K. pneumoniae MGH 78578 using paraquat and the corresponding oxidative stress regulon recorded using RNA-seq. The soxS gene was significantly induced during oxidative stress and a knock-out mutant was constructed, to explore its functionality. The wild-type and mutant were grown in the presence of paraquat and subjected to RNA-seq to elucidate the soxS regulon in K. pneumoniae MGH78578. Genes that are commonly regulated both in the oxidative stress regulon and soxS regulon were identified and denoted as the oxidative SoxS regulon - these included a stringent group of genes specifically regulated by SoxS. Efflux pump encoding genes such as acrAB-tolC, acrE, and global regulators such as marRAB were identified as part of this regulon. Consequently, the isogenic soxS mutant was found to exhibit a reduction in the minimum bactericidal concentration against tetracycline compared to that of the wild type. Impaired efflux activity, allowing tetracycline to be accumulated in the cytoplasm to bactericidal levels, was further evaluated using a tetraphenylphosphonium (TPP+) accumulation assay. The soxS mutant was also susceptible to tetracycline in vivo, in a zebrafish embryo model. We conclude that the soxS gene could be considered as a genetic target against which an inhibitor could be developed in the future and used in combinatorial therapy with tetracycline to combat infections associated with multi-drug resistant K. pneumoniae.

microbiology↗

bioRxiv2019

Yersinia canariae sp. nov., isolated from a human yersiniosis case

A Gram-negative rod from the Yersinia genus was isolated from a clinical case of yersiniosis in the United Kingdom. Long read sequencing data from an Oxford Nanopore Technology (ONT) MinION in conjunction with Illumina HiSeq reads were used to generate a finished quality genome of this strain. Overall Genome Related Index (OGRI) of the strain was used to determine that it was a novel species within Yersinia, despite biochemical similarities to Yersinia enterocolitica. The 16S ribosomal RNA gene accessions are MN434982-MN434987 and the accession number for the complete and closed chromosome is CP043727. The type strain is CFS3336T (=NCTC 14382T/ =LMG Accession under process).

microbiology↗