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Mechanisms of Resistance

Isolation of Nitrofurantoin-Resistant Mutants of Nitroreductase-Producing Clostridium sp. Strains from the Human Intestinal Tract

Fatemeh Rafii, Eugene B. Hansen Jr.
Fatemeh Rafii
Division of Microbiology and Chemistry, National Center for Toxicological Research, Food and Drug Administration, Jefferson, Arkansas 72079
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Eugene B. Hansen Jr.
Division of Microbiology and Chemistry, National Center for Toxicological Research, Food and Drug Administration, Jefferson, Arkansas 72079
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DOI: 10.1128/AAC.42.5.1121
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    Fig. 1.

    Effect of the concentration of nitrofurantoin on the growth of resistant strains of Clostridium species. (A)C. perfringens 3626NF; (B) C. paraputrificum NR1NF; (C) Clostridium sp. strain NR8NF. ○, no drug; •, 10 μg of nitrofurantoin per ml; □, 20 μg of nitrofurantoin per ml; ▪, 50 μg of nitrofurantoin per ml.

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    Fig. 2.

    HPLC elution profiles of cultures of a nitrofurantoin-resistant mutant, Clostridium sp. strain NR9NF, in BHI medium containing nitrofurantoin. (A) Culture inoculated with the mutant and sampled at zero time. (B) Culture inoculated with the mutant after incubation. (The elution profile of the noninoculated medium with nitrofurantoin after incubation for 24 h and that of nitrofurantoin after incubation with a heat-treated culture supernatant were both similar to that shown in panel A; the profile of the nitrofurantoin after incubation with an active culture supernatant was similar to that shown in panel B.)

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    Fig. 3.

    Areas of the nitrofurantoin peaks in HPLC profile after anaerobic incubation with overnight cultures of Clostridiumsp. mutants or noninoculated BHI medium. ▪, C. paraputrificum NR1NF; ▨, C. perfringens3626NF; ▤, Clostridium sp. strain NR8NF; ▨, C. leptum NP15NF; □,Clostridium sp. strain NR9NF; ▩, BHI medium.

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    Fig. 4.

    (A) Number of surviving cells of a nitrofurantoin-sensitive bacterium (Bacillus sp. strain PS) before and after incubation with either BHI broth or cultures of nitrofurantoin-resistant strains of Clostridium species (grown with or without nitrofurantoin). ▪, BHI broth; □, BHI and nitrofurantoin; •, C. paraputrificum NR1; ○, C. paraputrificum NR1NF with nitrofurantoin; ✚,Clostridium sp. strain NR9; ×, Clostridium sp. strain NR9NF grown with nitrofurantoin. (B) Number of surviving cells of a nitrofurantoin-sensitive bacterium (Bacillus sp. strain PS) before and after incubation with either BHI broth or cultures of nitrofurantoin-resistant strains ofClostridium species (grown with or without nitrofurantoin). ▴, BHI broth; ▵, BHI and nitrofurantoin; ▪, C. perfringens ATCC 3626; □, C. perfringens3626NF with nitrofurantoin; ✚, C. leptum NP15; ×, C. leptum NP15NF grown with nitrofurantoin; ○, Clostridium sp. strain NR8; •, Clostridiumsp. strain NR8NF. The incubation of Bacillus sp. strain PS with the supernatants of nitrofurantoin-resistant bacteria in the absence of nitrofurantoin produced results identical to those obtained for parental strains (data not shown).

Tables

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  • Table 1.

    Nitroreductase activity in cultures and cell-free supernatants of parental strains and nitrofurantoin-resistant mutants of Clostridium spp.

    StrainNitroreductase activity fora:
    CellsSoluble protein
    Parental strainResistant strainParental strainResistant strain
    Clostridium perfringens ATCC 362689.5102.66.810.4
    Clostridium paraputrificum NR139.8135.41.12.8
    Clostridium sp. strain NR874.7102.32.73.3
    Clostridium sp. strain NR9156.1227.03.33.9
    Clostridium leptumNP15NDND5.421.8
    • ↵a Results are expressed as units of activity where a unit is the amount of enzyme that produces 1 μg of 4-aminobenzoic acid per h from 4-nitrobenzoic acid at 37°C: for cells, units of activity per cell absorbance (A600); for protein, units of activity per milligram of soluble protein. ND, not determined.

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Isolation of Nitrofurantoin-Resistant Mutants of Nitroreductase-Producing Clostridium sp. Strains from the Human Intestinal Tract
Fatemeh Rafii, Eugene B. Hansen Jr.
Antimicrobial Agents and Chemotherapy May 1998, 42 (5) 1121-1126; DOI: 10.1128/AAC.42.5.1121

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Isolation of Nitrofurantoin-Resistant Mutants of Nitroreductase-Producing Clostridium sp. Strains from the Human Intestinal Tract
Fatemeh Rafii, Eugene B. Hansen Jr.
Antimicrobial Agents and Chemotherapy May 1998, 42 (5) 1121-1126; DOI: 10.1128/AAC.42.5.1121
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KEYWORDS

Bacterial Proteins
Clostridium
Nitroreductases

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