scholarly journals Interplay between antibiotic efficacy and drug-induced lysis underlies enhanced biofilm formation at subinhibitory drug concentrations

2017 ◽  
Author(s):  
Wen Yu ◽  
Kelsey Hallinen ◽  
Kevin B. Wood
Keyword(s):  
Cell Wall ◽  
Biofilm Formation ◽  
Cell Lysis ◽  
Living Cells ◽  
Cell Wall Synthesis ◽  
Drug Induced ◽  
Trade Off ◽  
Drug Concentrations ◽  
Subinhibitory Concentrations ◽  
Antibiotic Efficacy

Subinhibitory concentrations of antibiotics have been shown to enhance biofilm formation in multiple bacterial species. While antibiotic exposure has been associated with modulated expression in many biofilm-related genes, the mechanisms of drug-induced biofilm formation remain a focus of ongoing research efforts and may vary significantly across species. In this work, we investigate antibiotic-induced biofilm formation inE. faecalis, a leading cause of nosocomial infections. We show that biofilm formation is enhanced by subinhibitory concentrations of cell wall synthesis inhibitors, but not by inhibitors of protein, DNA, folic acid, or RNA synthesis. Furthermore, enhanced biofilm is associated with increased cell lysis, an increase in extracellular DNA (eDNA), and an increase in the density of living cells in the biofilm. In addition, we observe similar enhancement of biofilm formation when cells are treated with non-antibiotic surfactants that induce cell lysis. These findings suggest that antibiotic-induced biofilm formation is governed by a trade-off between drug toxicity and the beneficial effects of cell lysis. To understand this trade-off, we developed a simple mathematical model that predicts changes to antibiotic-induced biofilm formation due to external perturbations, and we verify these predictions experimentally. Specifically, we demonstrate that perturbations that reduce eDNA (DNase treatment) or decrease the number of living cells in the planktonic phase (a second antibiotic) decrease biofilm induction, while chemical inhibitors of cell lysis increase relative biofilm induction and shift the peak to higher antibiotic concentrations. Overall, our results offer experimental evidence linking cell wall synthesis inhibitors, cell lysis, increased eDNA, and biofilm formation inE. faecaliswhile also providing a predictive, quantitative model that sheds light on the interplay between cell lysis and antibiotic efficacy in developing biofilms.

scholarly journals Interplay between Antibiotic Efficacy and Drug-Induced Lysis Underlies Enhanced Biofilm Formation at Subinhibitory Drug Concentrations

2017 ◽  
Vol 62 (1) ◽  
Author(s):  
Wen Yu ◽  
Kelsey M. Hallinen ◽  
Kevin B. Wood
Keyword(s):  
Cell Wall ◽  
Biofilm Formation ◽  
Cell Lysis ◽  
Living Cells ◽  
Cell Wall Synthesis ◽  
Drug Induced ◽  
Trade Off ◽  
Content Type ◽  
Subinhibitory Concentrations ◽  
Antibiotic Efficacy

ABSTRACTSubinhibitory concentrations of antibiotics have been shown to enhance biofilm formation in multiple bacterial species. While antibiotic exposure has been associated with modulated expression of many biofilm-related genes, the mechanisms of drug-induced biofilm formation remain a focus of ongoing research efforts and may vary significantly across species. In this work, we investigate antibiotic-induced biofilm formation inEnterococcus faecalis, a leading cause of nosocomial infections. We show that biofilm formation is enhanced by subinhibitory concentrations of cell wall synthesis inhibitors but not by inhibitors of protein, DNA, folic acid, or RNA synthesis. Furthermore, enhanced biofilm is associated with increased cell lysis, increases in extracellular DNA (eDNA) levels, and increases in the density of living cells in the biofilm. In addition, we observe similar enhancement of biofilm formation when cells are treated with nonantibiotic surfactants that induce cell lysis. These findings suggest that antibiotic-induced biofilm formation is governed by a trade-off between drug toxicity and the beneficial effects of cell lysis. To understand this trade-off, we developed a simple mathematical model that predicts changes in antibiotic-induced biofilm formation due to external perturbations, and we verified these predictions experimentally. Specifically, we demonstrate that perturbations that reduce eDNA (DNase treatment) or decrease the number of living cells in the planktonic phase (a second antibiotic) decrease biofilm induction, while chemical inhibitors of cell lysis increase relative biofilm induction and shift the peak to higher antibiotic concentrations. Overall, our results offer experimental evidence linking cell wall synthesis inhibitors, cell lysis, increased eDNA levels, and biofilm formation inE. faecaliswhile also providing a predictive quantitative model that sheds light on the interplay between cell lysis and antibiotic efficacy in developing biofilms.

scholarly journals Copper(II) and Zinc(II) Complexes with the Clinically Used Fluconazole: Comparison of Antifungal Activity and Therapeutic Potential

2020 ◽  
Vol 14 (1) ◽  
pp. 24
Author(s):  
Nevena Lj. Stevanović ◽  
Ivana Aleksic ◽  
Jakob Kljun ◽  
Sanja Skaro Bogojevic ◽  
Aleksandar Veselinovic ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Biofilm Formation ◽  
Candida Parapsilosis ◽  
Therapeutic Potential ◽  
A549 Cells ◽  
Candida Krusei ◽  
Strong Inhibition ◽  
Subinhibitory Concentrations ◽  
Pyocyanin Production

Copper(II) and zinc(II) complexes with clinically used antifungal drug fluconazole (fcz), {[CuCl2(fcz)2].5H2O}n, 1, and {[ZnCl2(fcz)2]·2C2H5OH}n, 2, were prepared and characterized by spectroscopic and crystallographic methods. The polymeric structure of the complexes comprises four fluconazole molecules monodentately coordinated via the triazole nitrogen and two chlorido ligands. With respect to fluconazole, complex 2 showed significantly higher antifungal activity against Candida krusei and Candida parapsilosis. All tested compounds reduced the total amount of ergosterol at subinhibitory concentrations, indicating that the mode of activity of fluconazole was retained within the complexes, which was corroborated via molecular docking with cytochrome P450 sterol 14α-demethylase (CYP51) as a target. Electrostatic, steric and internal energy interactions between the complexes and enzyme showed that 2 has higher binding potency to this target. Both complexes showed strong inhibition of C. albicans filamentation and biofilm formation at subinhibitory concentrations, with 2 being able to reduce the adherence of C. albicans to A549 cells in vitro. Complex 2 was able to reduce pyocyanin production in Pseudomonas aeruginosa between 10% and 25% and to inhibit its biofilm formation by 20% in comparison to the untreated control. These results suggest that complex 2 may be further examined in the mixed Candida-P. aeruginosa infections.

scholarly journals The Effect of Hydroxylysine on Cell Wall Synthesis and Cell Stability in Streptococcus faecalis

1962 ◽  
Vol 237 (4) ◽  
pp. 1198-1202
Author(s):  
W. Grady Smith ◽  
Mary Newman ◽  
Franklin R. Leach ◽  
L.M. Henderson
Keyword(s):  
Cell Wall ◽  
Cell Wall Synthesis ◽  
Streptococcus Faecalis ◽  
Cell Stability

scholarly journals Direct Interaction of Polar Scaffolding Protein Wag31 with Nucleoid-Associated Protein Rv3852 Regulates Its Polar Localization

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1558
Author(s):  
Rajni Garg ◽  
Chinmay Anand ◽  
Sohini Ganguly ◽  
Sandhya Rao ◽  
Rinkee Verma ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Envelope ◽  
Transmembrane Helix ◽  
Ectopic Expression ◽  
Direct Interaction ◽  
Fine Tuning ◽  
Scaffolding Protein ◽  
Physical Interaction ◽  
Cell Wall Synthesis ◽  
Interaction Approach

Rv3852 is a unique nucleoid-associated protein (NAP) found exclusively in Mycobacterium tuberculosis (Mtb) and closely related species. Although annotated as H-NS, we showed previously that it is very different from H-NS in its properties and is distinct from other NAPs, anchoring to cell membrane by virtue of possessing a C-terminal transmembrane helix. Here, we investigated the role of Rv3852 in Mtb in organizing architecture or synthesis machinery of cell wall by protein–protein interaction approach. We demonstrated a direct physical interaction of Rv3852 with Wag31, an important cell shape and cell wall integrity determinant essential in Mtb. Wag31 localizes to the cell poles and possibly acts as a scaffold for cell wall synthesis proteins, resulting in polar cell growth in Mtb. Ectopic expression of Rv3852 in M. smegmatis resulted in its interaction with Wag31 orthologue DivIVAMsm. Binding of the NAP to Wag31 appears to be necessary for fine-tuning Wag31 localization to the cell poles, enabling complex cell wall synthesis in Mtb. In Rv3852 knockout background, Wag31 is mislocalized resulting in disturbed nascent peptidoglycan synthesis, suggesting that the NAP acts as a driver for localization of Wag31 to the cell poles. While this novel association between these two proteins presents one of the mechanisms to structure the elaborate multi-layered cell envelope of Mtb, it also exemplifies a new function for a NAP in mycobacteria.

scholarly journals A Novel Nanobody Precisely Visualizes Phosphorylated Histone H2AX in Living Cancer Cells under Drug-Induced Replication Stress

Cancers ◽  
2021 ◽  
Vol 13 (13) ◽  
pp. 3317
Author(s):  
Eric Moeglin ◽  
Dominique Desplancq ◽  
Audrey Stoessel ◽  
Christian Massute ◽  
Jeremy Ranniger ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Mammalian Cells ◽  
Chromatin Modification ◽  
Replication Stress ◽  
Living Cells ◽  
Single Step ◽  
Dna Double Strand Breaks ◽  
Drug Induced ◽  
Histone H2ax ◽  
C Terminus

Histone H2AX phosphorylated at serine 139 (γ-H2AX) is a hallmark of DNA damage, signaling the presence of DNA double-strand breaks and global replication stress in mammalian cells. While γ-H2AX can be visualized with antibodies in fixed cells, its detection in living cells was so far not possible. Here, we used immune libraries and phage display to isolate nanobodies that specifically bind to γ-H2AX. We solved the crystal structure of the most soluble nanobody in complex with the phosphopeptide corresponding to the C-terminus of γ-H2AX and show the atomic constituents behind its specificity. We engineered a bivalent version of this nanobody and show that bivalency is essential to quantitatively visualize γ-H2AX in fixed drug-treated cells. After labelling with a chemical fluorophore, we were able to detect γ-H2AX in a single-step assay with the same sensitivity as with validated antibodies. Moreover, we produced fluorescent nanobody-dTomato fusion proteins and applied a transduction strategy to visualize with precision γ-H2AX foci present in intact living cells following drug treatment. Together, this novel tool allows performing fast screenings of genotoxic drugs and enables to study the dynamics of this particular chromatin modification in individual cancer cells under a variety of conditions.

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