Fluorometric determination of ethidium bromide efflux kinetics in Escherichia coli

Paixão, Laura; Rodrigues, Liliana; Couto, Isabel; Martins, Marta; Fernandes, Pedro; Carvalho, Carla C. C. R. de; Monteiro, Gabriel A.; Sansonetty, Filipe; Amaral, Leonard; Viveiros, Miguel

doi:10.1186/1754-1611-3-18

Cited by 157 publications

(158 citation statements)

References 25 publications

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“…Diversos autores relacionam a resistência bacteriana aos desinfetantes à presença de mecanismos como o eϐluxo de substâncias pelas células (Paixão et al 2009, Kenneth et al 2009, Srinivasan et al 2009). Os resultados aqui encontrados reforçam esta hipótese, uma vez que os isolados mostraram-se muito sensíveis aos desinfetantes e o mecanismo de eϐluxo não foi observado.…”

Section: Discussionunclassified

Suscetibilidade a desinfetantes e perfil de resistência a antimicrobianos em isolados de Escherichia coli

et al. 2012

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RESUMO.-A colibacilose, causada por Escherichia coli, é a enfermidade entérica de maior impacto na produção de suínos, podendo levar à morte do animal. Esta bactéria possui grande capacidade de desenvolver resistência a múltiplos antimicrobianos e a desinfetantes. Desta forma, estudos que abordem mecanismos de resistência e perϐil de amostras de campo tornam-se necessários. E. coli é amplamente utilizada como modelo de estudos que exploram a resistência intrín-seca e extrínseca a multidrogas. Neste trabalho, buscou-se Colibacillosis caused by Escherichia coli is the most important enteric disease in pig production, which may lead to death of the affected animal. The bacterium has a great ability to develop resistance to multiple antibiotics and disinfectants. Thus, investigation addressing mechanisms of resistance and proϐile of ϐield samples is necessary. E. coli is widely used as a model for studies that explore the intrinsic and extrinsic resistance to multidrugs. In this paper, we attempt to associate the susceptibility proϐile of 62 isolates of E. coli to three disinfectants and 13 antimicrobials. Also 31 isolates were tested for the presence of efϐlux mechanism. Of the three disinfectants tested, alkyldimethylbenzylammonium chloride+nonyl phenoxy polyethoxy ethanol was the most effective (100%), followed by glutaraldehyde+al kyldimethylbenzylammonium chloride (95.2%) and alkyldimethylbenzylammonium chloride (88.8%). Among the antimicrobials tested, there was greater resistance to tetracycline (62.2%) and higher sensitivity to ϐlorfenicol (88.6%). The high sensitivity of the isolates against disinfectants may be related to the absence of efϐlux mechanism. The average index of multiple resistance to antimicrobials was 0.52, what demonstrates a proϐile of multidrug resistant isolates, showing the need for rational use of these drugs in pig production.

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Section: Discussionunclassified

Suscetibilidade a desinfetantes e perfil de resistência a antimicrobianos em isolados de Escherichia coli

et al. 2012

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“…To evaluate the activity of EPs, Paixão et al (2009) chose EtBr as the substrate for efflux investigation. EtBr emits weak fluorescence, but when it enters cells and combines with DNA, it emits strong fluorescence.…”

Section: Discussionmentioning

confidence: 99%

“…Hence, the bacteria become insensitive to antibiotics. Several methods have been used to measure EP system activity, such as an ethidium bromide (EtBr) accumulation assay (Paixão et al, 2009), an EP inhibition assay (Chollet et al, 2004), radiolabelled antibiotics (Hasdemir et al, 2004), and other methods (Daugelavičius et al, 2010). The most commonly used method is the EtBr accumulation assay, which is based on monitoring the accumulation of the fluorescent probe, EtBr (Paixão et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Influence of induced ciprofloxacin resistance on efflux pump activity of Klebsiella pneumoniae

Zhong

Zhang

Pan

et al. 2013

J. Zhejiang Univ. Sci. B

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Abstract:Objective: The efflux pump (EP) is one of the major mechanisms of antibiotic resistance in Klebsiella pneumoniae. However, there are few reports on the effect of the abuse of antibiotic use on the activity of EPs. To determine whether the use of low efficacy antibiotics has any effect on the activity of EPs and induces drug resistance in K. pneumoniae, we investigated the effect of ciprofloxacin on the activity of EPs in K. pneumoniae strains. Methods: Sixteen susceptible K. pneumoniae strains were isolated from patients and their minimum inhibitory concentrations (MICs) of ciprofloxacin were measured in the absence and presence of the pump inhibitor carbonyl cyanide m-chlorophenyl hydrazone (CCCP). The strains were then induced with a gradient of ciprofloxacin until the MICs of the strains showed no further increase, to obtain induced resistant strains. The EP activities of the strains before and after induction were compared using EP inhibition and ethidium bromide (EtBr) accumulation assays. Results: The MIC values of the strains were 16-256 times higher after induction than before induction. In the presence of CCCP, the MIC values of 50% of the induced strains were 2-4-fold lower than that in the absence of this inhibitor. The EtBr accumulation assay showed that the fluorescence of EtBr in the induced cells was lower than that in the cells before induction. Conclusions: EPs are widespread in susceptible and drug-resistant K. pneumoniae strains. Induction with ciprofloxacin may increase the activity of EPs in K. pneumoniae. The EtBr accumulation assay is more sensitive than the EP inhibition assay in evaluating the activity of EPs in K. pneumoniae.

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“…Membrane-disruptive activity of Amoxicillin, diclofenac sodium, aspirin, indomethacin, ibuprofen and amoxicillin/aspirin on in vitro grown E. coli (ATCC 8739) was determined by measuring the fluorescence enhancement of ethidium bromide (Sigma) (Paixão et al, 2009). To this end, E. coli were grown in tryptone broth medium in the presence or absence of amoxicillin as positive control, NSAIDs and Amoxicillin combined with aspirin for 24 h. The bacterial culture was incubated with ethidium bromide for 20 min at room temperature in the dark.…”

Section: Membrane-permeability Assaymentioning

confidence: 99%

Evaluation of antibacterial activity of some non-steroidal anti-inflammatory drugs against Escherichia coli causing urinary tract infection

Ahmed¹,

El-Baky²,

Ahmed³

et al. 2016

Afr. J. Microbiol. Res.

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Extensive use of antibiotics for urinary tract infections has led to the emergence of drug-resistant microorganisms and one solution to this problem is to search for non-antibiotic compounds that exert anti-bacterial activity through different mechanisms such as non-steroidal anti-inflammatory drugs (NSAIDs). In this study, out of 100 urine samples; 48 Escherichia coli strains were detected, 47.9% were multi-drug resistant. The antibiogram resistance pattern of the strains was carried out by agar dilution method. Diclofenac sodium, indomethacin, aspirin and ibuprofen were tested against the E. coli isolates. Diclofenac sodium showed the lowest MIC 50 and MIC 90 ; 8 and 256 g/ml, respectively. Aspirin showed MIC 50 of 64 g/ml, while both indomethacin and ibuprofen showed MIC 50 of 256 g/ml. Indomethacin, aspirin and ibuprofen showed the same MIC 90 of 1024 g/ml. The combined effects of the four NSAIDs and five antibiotics (Amoxicillin, Augmentin, Cefotaxime, Ciprofloxacin and Gentamicin) were tested on five resistant clinical E. coli strains by checkerboard dilution technique. All the tested NSAIDs significantly reduced the minimum inhibitory concentrations (MICs) of antibiotics against the tested bacteria and fractional inhibitory concentration indices (FICIs) for this combination ranged from 0.03 to 0.5. In this study, leakage of intracellular components suggests that the effect of NSAIDs on E. coli could be the formation of pores in the plasma membrane and scanning electron microscopy (SEM) observations confirmed the damage to the structural integrity of the tested bacteria. In conclusion, NSAIDs showed antibacterial activity against E. coli causing urinary tract infections (UTIs), a combination of them and antibiotics exhibited good synergism and the mechanism of their action was by damaging the bacterial cell membrane.

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Fluorometric determination of ethidium bromide efflux kinetics in Escherichia coli

Cited by 157 publications

References 25 publications

Suscetibilidade a desinfetantes e perfil de resistência a antimicrobianos em isolados de Escherichia coli

Suscetibilidade a desinfetantes e perfil de resistência a antimicrobianos em isolados de Escherichia coli

Influence of induced ciprofloxacin resistance on efflux pump activity of Klebsiella pneumoniae

Evaluation of antibacterial activity of some non-steroidal anti-inflammatory drugs against Escherichia coli causing urinary tract infection

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