title: Practical agar-based disk potentiation test for detection of fosfomycin-nonsusceptible Escherichia coli clinical isolates producing glutathione S-transferases.
abstract: The number of reports concerning Escherichia coli clinical isolates that produce glutathione S-transferases responsible for fosfomycin resistance (FR-GSTs) has been increasing. We have developed a disk-based potentiation test in which FR-GST producers expand the growth inhibition zone around a Kirby-Bauer disk containing fosfomycin in combination with sodium phosphonoformate (PPF). PPF, an analog of fosfomycin, is a transition-state inhibitor of FosA(PA), a type of FR-GST from Pseudomonas aeruginosa. Considering its mechanism of action, PPF was expected to inhibit a variety of FR-GSTs. In the presence of PPF, zone enlargement around the disk containing fosfomycin was observed for FosA3-, FosA4-, and FosC2-producing E. coli clinical isolates. Moreover, the growth inhibition zone was remarkably enlarged when the Mueller-Hinton (MH) agar plate contained 25 μg/ml glucose-6-phosphate (G6P). When we retrospectively tested 12 fosfomycin-resistant (MIC, ≥256 μg/ml) E. coli clinical isolates from our hospital with the potentiation test, 6 FR-GST producers were positive phenotypically by potentiation disk and were positive for FR-GST genes: 5 harbored fosA3 and 1 harbored fosA4. To identify the production of FR-GSTs, we set the provisional cutoff value, 5-mm enlargement, by adding PPF to a fosfomycin disk on the MH agar plates containing G6P. Our disk-based potentiation test reliably identifies FR-GST producers and can be performed easily; therefore, it will be advantageous in epidemiological surveys and infection control of fosfomycin-resistant bacteria in clinical settings.
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Genki
authors.initials: G
authors.lastname: Nakamura
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan wachino@med.nagoya-u.ac.jp.
authors.forename: Jun-Ichi
authors.initials: J
authors.lastname: Wachino
authors.affiliation: Department of Pathophysiological Laboratory Sciences, Nagoya University Graduate School of Medicine, Higashi-ku, Nagoya, Aichi, Japan.
authors.forename: Natsumi
authors.initials: N
authors.lastname: Sato
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Kouji
authors.initials: K
authors.lastname: Kimura
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Yamada
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Wanchun
authors.initials: W
authors.lastname: Jin
authors.affiliation: Department of Bacteriology II, National Institute of Infectious Diseases, Musashi-Murayama, Tokyo, Japan.
authors.forename: Keigo
authors.initials: K
authors.lastname: Shibayama
authors.affiliation: Department of Infectious Diseases, Nagoya University Hospital, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Yagi
authors.affiliation: Department of Pathophysiological Laboratory Sciences, Nagoya University Graduate School of Medicine, Higashi-ku, Nagoya, Aichi, Japan.
authors.forename: Kumiko
authors.initials: K
authors.lastname: Kawamura
authors.affiliation: Department of Bacteriology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, Japan.
authors.forename: Yoshichika
authors.initials: Y
authors.lastname: Arakawa
mesh_terms: D000362:Agar; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D003470:Culture Media / Q000737:chemistry; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004361:Drug Tolerance; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology* / Q000302:isolation & purification; D004927:Escherichia coli Infections / Q000382:microbiology*; D017245:Foscarnet / Q000378:metabolism; D005578:Fosfomycin / Q000494:pharmacology*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa; D017422:Sequence Analysis, DNA
chemical_list: D000900:Anti-Bacterial Agents; D003470:Culture Media; D004269:DNA, Bacterial; D005578:Fosfomycin; D017245:Foscarnet; D000362:Agar; D005982:Glutathione Transferase