Single Cell Killing Kinetics Differentiate Phenotypic Bacterial Responses to Different Antibacterial Classes

Zhang, Yuewen; Kepiro, Ibolya E.; Ryadnov, Maxim G.; Pagliara, Stefano

doi:10.1128/spectrum.03667-22

Cited by 7 publications

(8 citation statements)

References 75 publications

(99 reference statements)

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“…The mother machine microfluidic device was engineered, assembled and used to conduct single cell time-lapse microscopy experiments as previously reported [ 31 , 42 , 49 ]. Briefly, overnight E. coli cultures were concentrated to an OD 600 of 50 through centrifugation and resuspension of the pellet in spent medium that was obtained by filtering the supernatant from the overnight culture twice.…”

Section: Methodsmentioning

confidence: 99%

Slow growing bacteria survive bacteriophage in isolation

Attrill,

Łapińska,

Westra

et al. 2023

ISME Communications

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The interactions between bacteria and bacteriophage have important roles in the global ecosystem; in turn changes in environmental parameters affect the interactions between bacteria and phage. However, there is a lack of knowledge on whether clonal bacterial populations harbour different phenotypes that respond to phage in distinct ways and whether the abundance of such phenotypes within bacterial populations is affected by variations in environmental parameters. Here we study the impact of variations in nutrient availability, bacterial growth rate and phage abundance on the interactions between the phage T4 and individual Escherichia coli cells confined in spatial refuges. Surprisingly, we found that fast growing bacteria survive together with all of their clonal kin cells, whereas slow growing bacteria survive in isolation. We also discovered that the number of bacteria that survive in isolation decreases at increasing phage doses possibly due to lysis inhibition in the presence of secondary adsorptions. We further show that these changes in the phenotypic composition of the E. coli population have important consequences on the bacterial and phage population dynamics and should therefore be considered when investigating bacteria-phage interactions in ecological, health or food production settings in structured environments.

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Section: Methodsmentioning

confidence: 99%

Slow growing bacteria survive bacteriophage in isolation

Attrill,

Łapińska,

Westra

et al. 2023

ISME Communications

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“…To investigate the hypothesis that tachyplesin causes less membrane damage to low accumulators, we employed our microfluidics-based microscopy platform to simultaneously measure tachyplesin-NBD accumulation and bacterial membrane integrity via propidium iodide (PI) 43 . We found a strong positive semi-log correlation between tachyplesin-NBD fluorescence and PI fluorescence (r 2 = 0.70, p-value < 0.0001): all bacteria exhibiting PI staining had a tachyplesin-NBD fluorescence greater than 800 a.u.…”

Section: Resultsmentioning

confidence: 99%

Heterogeneous efflux pump expression underpins phenotypic resistance to antimicrobial peptides

Lee,

Łapińska,

Tolle

et al. 2024

Preprint

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Antimicrobial resistance threatens the viability of modern medical interventions. There is a dire need of developing novel approaches to counter resistance mechanisms employed by starved or slow-growing pathogens that are refractory to conventional antimicrobial therapies. Antimicrobial peptides have been advocated as potential therapeutic solutions due to low levels of genetic resistance observed in bacteria against these compounds. However, here we show that subpopulations of stationary phase Escherichia coli and Pseudomonas aeruginosa survive tachyplesin treatment without genetic mutations. These phenotypic variants induce efflux, outer membrane vesicles secretion and membrane modifications in response to tachyplesin exposure, sequestering the peptide in their membranes where it cannot exert its antimicrobial activity. We discovered that formation of these phenotypic variants could be prevented by administering tachyplesin in combination with sertraline, an existing clinically used inhibitor of transmembrane transporters, suggesting a novel approach for combatting antimicrobial-refractory stationary phase bacteria.

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“…Next, we set out to test the hypothesis that phage-induced downregulation of efflux processes led to increased intracellular accumulation of ciprofloxacin during phage-ciprofloxacin combination therapy. We used our recently reported microfluidics-based time-lapse microscopy platform 57,58 and a fluorescent derivative of ciprofloxacin (i.e. ciprofloxacin-nitrobenzoxadiazole, henceforth ciprofloxacin-NBD) to measure intracellular accumulation of ciprofloxacin in individual stationary phase B. thailandensis cells.…”

Section: Resultsmentioning

confidence: 99%

“…The minimum inhibitory concentration (MIC) of each antibiotic employed was determined via the broth dilution method 79 . Briefly, each antibiotic was dissolved according to the manufacture's specifications and diluted in a two-fold dilution series in LB medium within wells of a 96-well plate.…”

Section: Determination Of Antibiotic Minimum Inhibitory Concentration...mentioning

confidence: 99%

Phage-induced efflux down-regulation boosts antibiotic efficacy

Kraus,

Łapińska,

Chawla

et al. 2023

Preprint

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The interactions between a virus and its host vary in space and time and are affected by the presence of molecules that alter the physiology of either the host or the virus. Determining the dynamics of these interactions is paramount for predicting the fate of bacterial and phage populations and for designing rational phage-antibiotic therapies. We study the interactions between stationary phase Burkholderia thailandensis and the phage phi Bp-AMP1. Although heterogeneous genetic resistance to phage rapidly emerges in B. thailandensis, the presence of phage enhances the efficacy of three major antibiotic classes, the quinolones, the beta-lactams and the tetracyclines, but antagonizes tetrahydrofolate synthesis inhibitors. Enhanced antibiotic efficacy is underpinned by reduced antibiotic efflux in the presence of phage. This new phage-antibiotic therapy allows for eradication of stationary phase bacteria, whilst requiring reduced antibiotic concentrations, which is crucial for treating infections in sites where it is difficult to achieve high antibiotic concentrations.

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Single Cell Killing Kinetics Differentiate Phenotypic Bacterial Responses to Different Antibacterial Classes

Abstract: Antibiotics act against bacterial pathogens by inhibiting their growth or killing them directly. Different modes of action determine different antibacterial responses, whereas phenotypic differences in bacteria can challenge the efficacy of antibiotics.

Cited by 7 publications

References 75 publications

Slow growing bacteria survive bacteriophage in isolation

Slow growing bacteria survive bacteriophage in isolation

Heterogeneous efflux pump expression underpins phenotypic resistance to antimicrobial peptides

Phage-induced efflux down-regulation boosts antibiotic efficacy

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