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Antimicrobial Agents and Chemotherapy, August 2004, p. 3207-3208, Vol. 48, No. 8
0066-4804/04/$08.00+0     DOI: 10.1128/AAC.48.8.3207-3208.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

LETTER TO THE EDITOR

nimE Gene in a Metronidazole-Susceptible Veillonella sp. Strain


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LETTER
 
Metronidazole (MTZ) is widely used for the prophylaxis and the treatment of infections caused by anaerobes. These infections are mainly due to Bacteroides fragilis, but several reports identified Veillonella spp. as pathogens in severe infections (5, 7). Despite the extensive use of MTZ, resistance was maintained at a very low level for a long time and was first described for B. fragilis in 1978 (4). In contrast, an increasing level (3 to 5%) of B. fragilis group strains with decreased susceptibility to MTZ was observed (2). The presence of nitroimidazole (nim) resistance genes was first reported in 1989 (3), and five resistance determinants, nimA to nimE, were characterized either on plasmids or chromosomes (9). Only the location of nimE was not reported when the gene was described for B. fragilis group strains (9). More recently, the nimF gene, sharing 78% identity in sequence with nimD, has been reported in a B. fragilis group strain (S. Behrendtz, H. Fang, M. Hedberg, and C. Edlund, Abstr. 3rd World Congr. Anaerob. Bact. Infect., abstr. 6.003, 2003).

We have studied MTZ susceptibility and presence of nim genes for 116 clinical isolates of Veillonella spp. These strains were recovered either in pure culture or in aerobic-anaerobic polymicrobial cultures from various samples in 106 different patients hospitalized or consulting at the Montpellier University Hospital between March 1999 and July 2001. MTZ MICs were determined by the NCCLS reference agar dilution method (8). According to the NCCLS breakpoints, all the Veillonella sp. strains were susceptible to MTZ (Table 1). A PCR method using universal primers was used to detect nim genes after extraction of total genomic DNA according to the rapid procedure proposed by Trinh and Reysset (10). An amplification product was obtained for one isolate for which the MTZ MIC was 4 mg/liter. The 458-bp amplified fragment obtained was sequenced and gave the maximum identity (99.8%) with the nimE gene sequence of B. fragilis ARU 6881 (GenBank accession number AJ244018). To search for a possible chromosomal location of this nimE gene, DNA electrophoresis of I-CeuI restriction fragments, Southern blotting, and hybridization with a nim digoxigenin-labeled probe were performed (1). None of the I-CeuI-generated fragments hybridized with the probe. A Bacteroides strain known to harbor a chromosomal copy of the nim gene was used as a positive control. On the other hand, plasmid purification was performed by using a Quantum Prep plasmid miniprep kit (Bio-Rad Laboratories). Plasmid DNA was apparently not detectable when analyzed by agarose gel electrophoresis, but plasmid extraction yielded positive nim amplification. Taken together, these results suggested that nimE was probably located on a plasmid present in low copy number in the Veillonella strain, and this could be related to the low MTZ MIC observed for this strain.


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  		TABLE 1. Repartition of the MTZ MICs obtained by the agar dilution method for the 116 Veillonella sp. clinical isolates

nim genes were mainly described in B. fragilis group isolates, and nimA is the only one presently recovered in other anaerobic bacteria: Propionibacterium spp., Actinomyces odontolyticus, Prevotella bivia, and Clostridium bifermentans (6). We report the first detection of a nim gene in a Veillonella sp. It is also the first description of the nimE gene outside the B. fragilis group. As previously described for the genus Bacteroides, the presence of nim genes did not lead systematically to the expression of MTZ resistance, but the isolation of this Veillonella sp. strain extends the list of nim gene-harboring bacteria.


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Nucleotide sequence accession number.
 
A partial sequence of 448 bp for the Veillonella sp. nitroimidazole resistance protein (nimE) gene has been deposited in GenBank under accession number AY575922.


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ACKNOWLEDGMENTS
 
We thank Laurent Calvet for excellent technical assistance in MTZ MIC determinations.


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REFERENCES
 
    1
  1. Bonnet, R., H. Marchandin, C. Chanal, D. Sirot, R. Labia, C. De Champs, E. Jumas-Bilak, and J. Sirot. 2002. Chromosome-encoded class D ß-lactamase OXA-23 in Proteus mirabilis. Antimicrob. Agents Chemother. 46:2004-2006.[Abstract/Free Full Text]
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  2. Breuil, J., C. Burnat, O. Patey, and A. Dublanchet. 1989. Survey of Bacteroides fragilis susceptibility patterns in France. J. Antimicrob. Chemother. 24:69-75.[Abstract/Free Full Text]
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  3. Breuil, J., A. Dublanchet, N. Truffaut, and M. Sebald. 1989. Transferable 5-nitroimidazole resistance in the Bacteroides fragilis group. Plasmid 21:151-154.[CrossRef][Medline]
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  4. Ingham, H. R., S. Eaton, C. W. Venables, and P. C. Adams. 1978. Bacteroides fragilis resistant to metronidazole after long-term therapy. Lancet 1:214.
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  5. Liu, J. W., J. J. Wu, L. R. Wang, L. J. Teng, and T. C. Huang. 1998. Two fatal cases of Veillonella bacteremia. Eur. J. Clin. Microbiol. Infect. Dis. 17:62-64.[CrossRef][Medline]
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  6. Lubbe, M. M., K. Stanley, L. J. Chalkley. 1999. Prevalence of nim genes in anaerobic/facultative anaerobic bacteria isolated in South Africa. FEMS Microbiol. Lett. 172:79-83.[CrossRef][Medline]
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  7. Marchandin, H., H. Jean-Pierre, C. Carrière, F. Canovas, H. Darbas, and E. Jumas-Bilak. 2001. Prosthetic joint infection due to Veillonella dispar. Eur. J. Clin. Microbiol. Infect. Dis. 20:340-342.[CrossRef][Medline]
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  8. NCCLS. 2001. Methods for antimicrobial susceptibility testing of anaerobic bacteria, 5th ed. Approved standard M11-A5. NCCLS, Wayne, Pa.
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  9. Stubbs, S. L., J. S. Brazier, P. R. Talbot, and B. I. Duerden. 2000. PCR-restriction fragment length polymorphism analysis for identification of Bacteroides spp. and characterization of nitroimidazole resistance genes. J. Clin. Microbiol. 38:3209-3213.[Abstract/Free Full Text]
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  10. Trinh, S., and G. Reysset. 1996. Detection by PCR of the nim genes encoding 5-nitroimidazole resistance in Bacteroides spp. J. Clin. Microbiol. 34:2078-2084.[Abstract]
H. Marchandin*
H. Jean-Pierre
J. Campos
Laboratoire de Bactériologie
Hôpital Arnaud de Villeneuve
371 Avenue du Doyen Gaston Giraud
34295 Montpellier Cedex 5, France

L. Dubreuil
Laboratoire de Bactériologie
Faculté de Pharmacie
Lille, France

C. Teyssier
E. Jumas-Bilak
Faculté de Pharmacie
Montpellier, France

* Phone: 33 4 67 33 58 84 Fax: 33 4 67 33 58 93. E-mail: h-marchandin{at}chu-montpellier.fr.

Antimicrobial Agents and Chemotherapy, August 2004, p. 3207-3208, Vol. 48, No. 8
0066-4804/04/$08.00+0     DOI: 10.1128/AAC.48.8.3207-3208.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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  • Soki, J., Gal, M., Brazier, J. S., Rotimi, V. O., Urban, E., Nagy, E., Duerden, B. I. (2006). Molecular investigation of genetic elements contributing to metronidazole resistance in Bacteroides strains. J Antimicrob Chemother 57: 212-220 [Abstract] [Full Text]  

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