Servicio de Bacteriología, Laboratorio de Referencia de Meningococos, Centro Nacional de Microbiología, Instituto de Salud Carlos III, 28220 Majadahonda, Madrid, Spain
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Abstract |
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Introduction |
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The role of N. lactamica in the emergence of N. meningitidis strains with intermediate resistance to penicillin (Peni), by horizontal genetic exchange of penicillin binding protein (PBP) genes, has been proposed in several studies.2,3 Intragenic recombination of the meningococci chromosomal gene penA, encoding the target site for penicillin, with related genes of N. lactamica and other commensal Neisseria spp. may have generated mosaic genes that encode proteins with reduced affinity for penicillin, resulting in the emergence of Peni meningococcal strains.4
The aim of this study was to analyse the levels of susceptibility of N. lactamica isolates to antimicrobial agents commonly used for treatment and prophylaxis of MD.
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Material and methods |
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Susceptibility to penicillin, ampicillin, cefotaxime, ceftriaxone, rifampicin and ciprofloxacin was determined by the agar dilution method in MuellerHinton agar (Difco Laboratories, Detroit, MI, USA) supplemented with 5% sheep blood, with a final inoculum of 104105 cfu per spot. The doubling dilution ranges tested were 0.00716 mg/L for penicillin and ampicillin, 0.00031 mg/L for cefotaxime and ceftriaxone, 0.0072 mg/L for rifampicin and 0.00071 mg/L for ciprofloxacin.
Plates were incubated for 20 h at 37°C in air containing 5% CO2. The MIC was defined as the lowest concentration at which no growth was visible. The categories of susceptible, intermediate and resistant were defined according to the criteria used for N. meningitidis.4 Staphylococcus aureus ATCC 29213 and Escherichia coli ATCC 25922 were used as quality control organisms and were included each time isolates were tested.
In order to investigate the possible role of membrane permeability in the susceptibility of N. lactamica to rifampicin the MICs of this drug were determined in the presence and absence of Tween 80 (0.05%) for 41 isolates according to the protocol described previously.6
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Results and discussion |
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Most of the isolates were susceptible to rifampicin. MIC50 and MIC90 of rifampicin were higher than those described for N. meningitidis,7 although no isolate was resistant. Changes in the susceptibility to rifampicin may be because of alterations in membrane permeability,6 so differences in the membrane composition between N. meningitidis and N. lactamica might be the explanation for the differences in the susceptibility levels to rifampicin. Tween 80 is a detergent that has been used to investigate membrane permeability in N. meningitidis.6 In the study of Abadi et al.,6 no difference was observed with meningococcal strains with intermediate levels of resistance. However, we nonetheless decided to use Tween 80 for N. lactamica isolates with intermediate resistance to rifampicin because N. meningitidis might have a different membrane composition in comparison with N. lactamica. No difference in growth rates in the presence and absence of Tween 80 was observed for isolates with different MICs. Point mutations in the rpoB gene in N. lactamica could also account for the higher rifampicin MICs seen for N. lactamica compared with meningococci.7 Sequencing of this gene in N. lactamica and its comparison with the rpoB sequence in N. meningitidis (susceptible and resistant isolates) should be undertaken to resolve this issue.
Finally, six isolates showed intermediate susceptibility to ciprofloxacin, with MICs of 0.12 mg/L (n = 4) and 0.5 mg/L (n = 2). Although isolates with this level of susceptibility are frequent in N. gonorrhoeae,4 such isolates have not been described among those Neisseria species that share an ecological niche with N. meningitidis. Such N. lactamica strains might constitute a potential DNA source for the development of ciprofloxacin resistance in N. meningitidis. To date only two meningococcal strains with decreased susceptibility to ciprofloxacin have been described.10 Further studies will be necessary in order to determine whether these N. lactamica strains show changes in gyrA and parC genes similar to those described in gonococci. Transformation experiments with the DNA of N. lactamica strains with decreased susceptibility to ciprofloxacin and susceptible meningococci isolates could demonstrate whether horizontal spread of ciprofloxacin resistance between meningococci and N. lactamica can easily happen.
Horizontal genetic exchange of antimicrobial resistance determinants between commensal Neisseria species, such as N. lactamica and N. meningitidis is, perhaps, one of the most important mechanisms in the continuing evolution of resistance to antimicrobial agents in N. meningitidis.4
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Acknowledgements |
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Notes |
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References |
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2 . Sáez-Nieto, J. A., Luján, R., Matinez-Suarez, J. V., Berrón, S., Vázquez, J. A., Viñas, M. et al. (1990). Neisseria lactamica and Neisseria polysaccharea as possible sources of meningococcal ß-lactam resistance by genetic transformation. Antimicrobial Agents and Chemotherapy 34, 226972.[ISI][Medline]
3 . Luján, R., Zhang, Q. Y., Sáez-Nieto, J. A., Jones, D. M. & Spratt, B. G. (1991). Penicillin resistant isolates of Neisseria lactamica produced altered forms of penicillin binding protein 2 that arose by interspecies gene transfer. Antimicrobial Agents and Chemotherapy35, 3004.[ISI][Medline]
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6 . Abadi, F. J. R., Carter, P. E., Cash, P. & Pennington, T. H. (1996). Rifampin resistance in Neisseria meningitidis due to alterations in membrane permeability. Antimicrobial Agents and Chemotherapy 40, 64651.[Abstract]
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8 . Baquero, F., Martínez Beltrán, J. & Cantón, R. (1997). MENSURA break-points. Spanish criteria for determination of susceptibility to antibiotics. Revista Española de Quimioterapia 10, 30313.
9 . Sáez-Nieto, J. A., Luján, R., Berrón, S., Campos, J., Viñas, M., Fusté, C. et al. (1992). Epidemiology and molecular basis of penicillin-resistant Neisseria meningitidis in Spain: A 5-year history (19851989). Clinical Infectious Diseases 14, 394402.[ISI][Medline]
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Shultz, T. R., Tapsall, J. W., White, P. A. & Newton, P. J. (2000). An invasive isolate of Neisseria meningitidis showing decreased susceptibility to quinolones. Antimicrobial Agents and Chemotherapy 44, 1116.
Received 11 July 2001; returned 19 September 2001; revised 7 November 2001; accepted 29 November 2001