Urinary Tract Conditions Affect Fosfomycin Activity against Escherichia coli Strains Harboring Chromosomal Mutations Involved in Fosfomycin Uptake

Martín-Gutiérrez, Guillermo; Docobo-Pérez, Fernando; Rodríguez-Beltrán, Jerónimo; Rodríguez-Martínez, José Manuel; Aznar, Javier; Pascual, Álvaro; Blázquez, Jesús

doi:10.1128/aac.01899-17

Cited by 28 publications

(40 citation statements)

References 45 publications

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“…FOS is administered by both oral and i.v. routes (Martin-Gutierrez et al, 2018). In vitro (Kastoris, Rafailidis, Vouloumanou, Gkegkes, & Falagas, 2010;Samonis, Maraki, Karageorgopoulos, Vouloumanou, & Falagas, 2012) as well as prospective clinical (Patwardhan & Singh, 2017;Sardar, Basireddy, Navaz, Singh, & Kabra, 2017) studies have shown promising bactericidal activity of FOS (alone or in combination with other antibiotics) against various strains of urinary pathogens, including Gram-positive and Gramnegative bacteria.…”

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confidence: 99%

Development and validation of a LC‐MS/MS method for quantitation of fosfomycin – Application to in vitro antimicrobial resistance study using hollow‐fiber infection model

Gandhi

Matta

Garimella

et al. 2018

Biomedical Chromatography

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Extensive use and misuse of antibiotics over the past 50 years has contributed to the emergence and spread of antibiotic-resistant bacterial strains, rendering them as a global health concern. To address this issue, a dynamic in vitro hollow-fiber system, which mimics the in vivo environment more closely than the static model, was used to study the emergence of bacterial resistance of Escherichia coli against fosfomycin (FOS). To aid in this endeavor we developed and validated a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for quantitative analysis of FOS in lysogeny broth. FOS was resolved on a Kinetex HILIC (2.1 × 50 mm, 2.6 μm) column with 2 mm ammonium acetate (pH 4.76) and acetonitrile as mobile phase within 3 min. Multiple reaction monitoring was used to acquire data on a triple quadrupole mass spectrometer. The assay was linear from 1 to 1000 μg/mL. Inter- and intra-assay precision and accuracy were <15% and between ±85 and 115% respectively. No significant matrix effect was observed when corrected with the internal standard. FOS was stable for up to 24 h at room temperature, up to three freeze-thaw cycles and up to 24 h when stored at 4°C in the autosampler. In vitro experimental data were similar to the simulated plasma pharmacokinetic data, further confirming the appropriateness of the experimental design to quantitate antibiotics and study occurrence of antimicrobial resistance in real time. The validated LC-MS/MS assays for quantitative determination of FOS in lysogeny broth will help antimicrobial drug resistance studies.

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confidence: 99%

Development and validation of a LC‐MS/MS method for quantitation of fosfomycin – Application to in vitro antimicrobial resistance study using hollow‐fiber infection model

Gandhi

Matta

Garimella

et al. 2018

Biomedical Chromatography

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“…Our results highlight the importance that the modification of the activity of enzymes belonging to central metabolism may have in the susceptibility to antibiotics, as fosfomycin, that are not known to interact with such enzymes. The finding that fosfomycin activity is highly dependent on the bacterial metabolic status, being more active when bacteria grow intracellularly (27, 28) or under acidic conditions and anaerobiosis in urine (62), further support that antibiotic activity and, consequently antibiotic resistance, are interlinked with the bacterial metabolism.…”

Section: Discussionmentioning

confidence: 88%

The inactivation of enzymes belonging to the central carbon metabolism, a novel mechanism of developing antibiotic resistance

Gil-Gil

Corona

Martínez

et al. 2019

Preprint

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Fosfomycin is a bactericidal antibiotic, analogous to phosphoenolpyruvate (PEP) that exerts its activity by inhibiting the activity of MurA. This enzyme catalyzes the first step of peptidoglycan biosynthesis, the transfer of enolpyruvate from PEP to uridine-diphosphate-N-acetylglucosamine. Fosfomycin is increasingly used in the last years, mainly for treating infections caused by Gram-negative multidrug resistant bacteria as Stenotrophomonas maltophilia, an opportunistic pathogen characterized by its low susceptibility to antibiotics of common use. The mechanisms of mutational resistance to fosfomycin in S. maltophilia were studied in the current work. None of the mechanisms so far described for other organisms, which include the production of fosfomycin inactivating enzymes, target modification, induction of alternative peptidoglycan biosynthesis pathway and the impaired entrance of the antibiotic, are involved in the acquisition of such resistance by this bacterial species. Rather the unique cause of resistance in the studied mutants is the mutational inactivation of different enzymes belonging to the Embden-Meyerhof-Parnas central metabolism pathway. The amount of intracellular fosfomycin accumulation did not change in any of these mutants showing that neither the inactivation nor the transport of the antibiotic were involved. Transcriptomic analysis also showed that the mutants did not present changes in the expression level of putative alternative peptidoglycan biosynthesis pathway genes neither any related enzyme. Finally, the mutants did not present an increased PEP concentration that might compete with fosfomycin for its binding to MurA. Based on these results, we describe a completely novel mechanism of antibiotic resistance based on the remodeling of S. maltophilia metabolism.SignificanceAntibiotic resistance (AR) has been largely considered as a specific bacterial response to an antibiotic challenge. Indeed, its study has been mainly concentrated in mechanisms that affect the antibiotics (mutations in transporters, the activity of efflux pumps and antibiotic modifying enzymes) or their targets (i.e.: target mutations, protection or bypass). Usually, AR-associated metabolic changes were considered to be a consequence (fitness costs) and not a cause of AR. Herein, we show that strong alterations in the bacterial metabolism can also be the cause of AR. In the study here presented, Stenotrophomonas maltophilia acquires fosfomycin resistance through the inactivation of glycolytic enzymes belonging to the Embden-Meyerhof-Parnas. Besides resistance to fosfomycin, this inactivation also impairs the bacterial gluconeogenic pathway. Together with previous work showing that AR can be under metabolic control, our results provide evidence that AR is intertwined with the bacterial metabolism.

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“…In order to explain the unexpected activity of fosfomycin against both susceptible and resistant strains, investigations were performed to test the influence of acidic pH, as 85% of patients with UTI due to E. coli had a urine pH of < 6.5) (12). Indeed, pH in uninfected urine from 5 CBA mice before experimental pyelonephritis was 4.5 and ranged from 5.5 to 6.0 in the same mice after 48h of experimental pyelonephritis.…”

Section: Resultsmentioning

confidence: 99%

“…Fosfomycin activity observed in infected kidneys may result from the combination of local specific physiological conditions in urine and of fosfomycin concentrations in the kidneys. Indeed, several authors have already shown that low pH values increased fosfomycin in vitro activity (12, 19, 20). In an acidic environment such as urine, fosfomycin is partially protonated, being in a more lipophilic state, allowing fosfomycin entry into bacteria, and resulting in higher antimicrobial activity (20).…”

Section: Discussionmentioning

confidence: 97%

“…In addition, some mutations conferring fosfomycin resistance have been shown to decrease pilus biosynthesis and bacterial adhesion to epithelium cells (10, 11). Furthermore, Martin-Gutierrez et al recently demonstrated that anaerobiosis and acidic pH values in urine decreased fosfomycin MICs in strains harboring chromosomal resistance mutations (12). Finally, it has been previously shown that high concentrations (1000-4000 μg/ml) were achieved in urine after oral administration of 3 g of fosfomycin-tromethamine in humans and remained above 100 μg/ml for at least 30-40h (13), but data on concentrations in kidneys are scarce.…”

Section: Introductionmentioning

confidence: 99%

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Unexpected activity of oral fosfomycin against resistant strains ofEscherichia coliin murine pyelonephritis

Pourbaix

Guérin

Burdet

et al. 2019

Preprint

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26Fosfomycin-tromethamine activity is well established for oral treatment of uncomplicated 27 lower urinary tract infections but little is known about its potential efficacy in 28 pyelonephritis. Ascending pyelonephritis was induced in mice infected with 6 strains of 29 Escherichia coli (fosfomycin MICs: 1 µg/ml to 256 µg/ml). Urine pH was 4.5 before 30 infection and 5.5-6.0 during infection. Animals were treated for 24h with fosfomycin (100 31 mg/kg subcutaneously every 4 hours) and CFU were enumerated in kidneys 24h after the 32 last fosfomycin injection. Peak (20.5 µg/ml at 1h) and trough (3.5 µg/ml at 4h) levels in 33 plasma were comparable to those obtained in human after an oral dose of 3 grams. 34 Fosfomycin treatment significantly reduced bacterial loads in kidneys (3.65 log 10 CFU/g 35 [min-max=1.83-7.03] and 1.88 log 10 CFU/g [1.78-5.74] in start-of-treatment control mice 36 and treated mice, respectively, P < 10 -6 ). However, this effect was not found to differ 37 across the 6 study strains (P = 0.71) and between the 3 susceptible and the 3 resistant 38 strains (P=0.09). Three phenomena may contribute to explain this unexpected in vivo 39 activity: i) in mice, fosfomycin kidney/plasma concentrations ratio increased from 1 to 7.8 40 (95% CI, 5.2; 10.4) within 24 hours; in vitro, when pH decreased to 5: (ii) fosfomycin MICs 41 for the 3 resistant strains (64-256 µg/ml) decreased into the susceptible range (16-32 42 µg/ml) and: iii) maximal growth rates significantly decreased for all strains and were the 43 lowest in urine. These results suggest that local fosfomycin concentrations and 44 physiological conditions may favour fosfomycin activity in pyelonephritis, even against 45 resistant strains. 46 47 48

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Urinary Tract Conditions Affect Fosfomycin Activity against Escherichia coli Strains Harboring Chromosomal Mutations Involved in Fosfomycin Uptake

Cited by 28 publications

References 45 publications

Development and validation of a LC‐MS/MS method for quantitation of fosfomycin – Application to in vitro antimicrobial resistance study using hollow‐fiber infection model

Development and validation of a LC‐MS/MS method for quantitation of fosfomycin – Application to in vitro antimicrobial resistance study using hollow‐fiber infection model

The inactivation of enzymes belonging to the central carbon metabolism, a novel mechanism of developing antibiotic resistance

Unexpected activity of oral fosfomycin against resistant strains ofEscherichia coliin murine pyelonephritis

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