Morphological and ultrastructural changes in bacterial cells as an indicator of antibacterial mechanism of action

Cushnie, T.P. Tim; O’Driscoll, Noëlle H.; Lamb, Andrew J.

doi:10.1007/s00018-016-2302-2

Cited by 155 publications

(149 citation statements)

References 201 publications

(440 reference statements)

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“…Observed changes included cell bloating, a crushed appearance, cell surface disruption, and lysis. These cell alteration effects may be due to bioactive constituents in the culture filtrate extract disrupting peptidoglycan biosynthesis [23,24]. Many bioactive compounds produced by Streptomyces spp., for example phenolic compounds and antibiotics, have been shown to alter cell surface appearance.…”

Section: Discussionmentioning

confidence: 99%

Investigation of antibacterial and anti-cancer activities of Streptomyces sp SRF1 culture filtrate

Sangdee¹,

Buranrat²,

Seephonkai³

et al. 2018

Trop. J. Pharm Res

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Section: Discussionmentioning

confidence: 99%

Investigation of antibacterial and anti-cancer activities of Streptomyces sp SRF1 culture filtrate

Sangdee¹,

Buranrat²,

Seephonkai³

et al. 2018

Trop. J. Pharm Res

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“…12, the beading of the bacilli is succeeded by the formation of continuous filaments and then by aggregation, lysis and formation of necrotic spheroplasts, the transition to which increases with the duration of the treatment 54 . The inhibition of penicillin-binding protein 3, which crosslinks proteoglycan exclusively at the septal, but not the lateral wall, the inhibition of DNA synthesis through the SOS response and the inhibition of protein and/or RNA syntheses have all been invoked as possible downstream causes of bacterial filamentation 55 .…”

Section: Resultsmentioning

confidence: 99%

Antimicrobial hydroxyapatite–gelatin–silica composite pastes with tunable setting properties

Uskoković

Ghosh

2017

J. Mater. Chem. B

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Bone grafting is one of the commonest surgical procedures, yet all bone substitutes developed so far suffer from specific weaknesses and the search for a bone graft material with ideal physical and biological properties is still ongoing. Calcium phosphate pastes are the most frequently used synthetic bone grafts, yet they (a) often take an impractically long time to set, (b) release the drug content too fast, and (c) do not form pores large enough to accommodate host cells and foster osseointegration. To make up for these deficiencies, we introduced gelatin and silica to pastes composed of 5–15 nm sized hydroxyapatite nanoparticles and yielded a bioresorbable composite that is compact, yet flowing upon injection; that prevents setting at room temperature, but sets promptly, in minutes, at 37 °C; that displays an increase in surface porosity following immersion in physiological fluids; that allows for sustained release of antibiotics; and that sets in a tunable manner and in clinically relevant time windows: 1–3 minutes at its fastest. Timelapse, in situ X-ray diffraction analysis demonstrated that the setting process is accompanied by an increase in crystallinity of the initially amorphous hydroxyapatite, involving no polymorphic phase transitions in its course. Setting time can be tuned by controlling the weight content of gelatin or powder-to-liquid ratio. The release of vancomycin was slow, ~ 8 % after 2 weeks, and unaffected by the gelatin content. While vancomycin-loaded pastes were effective in reducing the concentration of all bacterial species analyzed, the bacteriostatic effects of the antibiotic-free pastes were pronounced against S. liquefaciens and E. coli. S. liquefaciens bacilli underwent beading and filamentation during the treatment, suggesting that the antimicrobial effects are attributable to cell wall disruption by hydroxyapatite nanoparticles. Vancomycin-loaded pastes augmented the activity of the antibiotic against P. aeruginosa and S. liquefaciens, while exhibiting no negative effects against human mesenchymal stem cells. They were also uptaken three times more abundantly than pure hydroxyapatite, indicating the theoretical favorability of their use for intracellular delivery of therapeutics. This selectivity, toxic for bacteria and harmless for primary stem cells, is promising for application as bone grafts for osteomyelitis.

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“…Filamentation was conspicuous by its absence in both species. Filamentation is a hallmark of DNA damage, induced via the SOS response by a diverse range of physical and chemical agents including UV, cosmomycin D, metronidazole, mitomycin C, the fluoroquinolones, novobiocin, and zidovudine (Cushnie et al 2016). The failure of colistin to induce filamentation at any concentration or time-point in either species argues strongly against the hypothesis that colistin kills bacterial cells through DNA damage or disruption of DNA synthesis.…”

Section: Discussionmentioning

confidence: 99%

“…A slightly higher bacterial cell density was needed in this assay compared to potassium loss and flow cytometry assays, as cells in untreated control populations of 1 × 10 6 cfu mL − 1 appear sparse when examined by SEM prior to 24 h incubation (data not shown) and antibiotic-induced morphological changes are known to occur much earlier than 24 h (sometimes within just 1 h of treatment; Cushnie et al 2016). The size of cells and cell aggregates was determined using LEO S430 software.…”

Section: Methodsmentioning

confidence: 99%

“…Cytoplasmic membrane damage was assessed by measuring potassium loss from colistin-treated cells, as leakage of this intracellular solute is an early sign that membrane integrity has been compromised (Yu et al 2016; Liang et al 2016). Colistin-treated bacteria were also examined by flow cytometry and scanning electron microscopy (SEM) as changes in the size and shape of cells can be useful in identifying antibiotic targets too (Peach et al 2013; Cushnie et al 2016). To account for the fact that antibiotic-induced morphological changes can vary with antibiotic concentration (Alsteens et al 2008), inoculum density (Diver and Wise 1986), and test bacterial species (Wojnicz et al 2007), we examined the effects of sub-inhibitory, inhibitory, and bactericidal concentrations of colistin upon two species, E. coli and P. aeruginosa , using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values determined for assay-specific inoculum densities.…”

Section: Introductionmentioning

confidence: 99%

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Colistin causes profound morphological alteration but minimal cytoplasmic membrane perforation in populations of Escherichia coli and Pseudomonas aeruginosa

et al. 2018

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Whilst colistin (polymyxin E) represents the last mainstream treatment option for multidrug-resistant Gram-negative pathogens, details of its mechanism of action remain to be fully resolved. In this study, the effects of sub-inhibitory, inhibitory-bactericidal, and supra-bactericidal levels of colistin on the membrane integrity and morphology of Escherichia coli and Pseudomonas aeruginosa were investigated using potassium loss, flow cytometry, and scanning electron microscopy (SEM). Supra-bactericidal colistin concentrations induced just 4–12% intracellular potassium loss from bacteria after 24 h. Flow cytometry data suggested colistin might alter cell arrangement, and SEM confirmed the antibiotic causes bacterial aggregation. Filamentation was not detected in either species at any concentration or time-point up to 24 h. These results argue against the hypotheses that colistin kills bacteria by puncturing the cytoplasmic membrane or disrupting DNA synthesis. The colistin-induced bacterial aggregation detected has implications for the interpretation of MBC, time-kill, and other test results obtained with this antibiotic.Electronic supplementary materialThe online version of this article (10.1007/s00203-018-1485-3) contains supplementary material, which is available to authorized users.

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Morphological and ultrastructural changes in bacterial cells as an indicator of antibacterial mechanism of action

Cited by 155 publications

References 201 publications

Investigation of antibacterial and anti-cancer activities of Streptomyces sp SRF1 culture filtrate

Investigation of antibacterial and anti-cancer activities of Streptomyces sp SRF1 culture filtrate

Antimicrobial hydroxyapatite–gelatin–silica composite pastes with tunable setting properties

Colistin causes profound morphological alteration but minimal cytoplasmic membrane perforation in populations of Escherichia coli and Pseudomonas aeruginosa

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